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 Bioinformatics
A Computing Perspective
 Bioinformatics
A Computing Perspective

Shuba Gopal
Rochester Institute of Technology

Anne Haake
Rochester Institute of Technology

Rhys Price Jones

George Washington University

Paul Tymann
Rochester Institute of Technology
BIOINFORMATICS: A COMPUTING PERSPECTIVE

Published by McGraw-Hill, a business unit of The McGraw-Hill Companies, Inc., 1221 Avenue of the Americas, New York,
NY 10020. Copyright © 2009 by The McGraw-Hill Companies, Inc. All rights reserved. No part of this publication may be
reproduced or distributed in any form or by any means, or stored in a database or retrieval system, without the prior written
consent of The McGraw-Hill Companies, Inc., including, but not limited to, in any network or other electronic storage or
transmission, or broadcast for distance learning.

Some ancillaries, including electronic and print components, may not be available to customers outside the United States.

This book is printed on acid-free paper.

1 2 3 4 5 6 7 8 9 0 DOC/DOC 0 9 8

ISBN 978–0–07–313364–5
MHID 0–07–313364–7

Global Publisher: Raghothaman Srinivasan

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Senior Project Manager: Kay J. Brimeyer
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Photo Credits: Tymann Bioinformatics 1/e

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Researchers, Inc.,

Library of Congress Cataloging-in-Publication Data

Bioinformatics : a computing perspective / Shuba Gopal ... [et al.]. – 1st ed.
p. cm.
Includes index.
ISBN 978–0–07–313364–5 — ISBN 0–07–313364–7 (hard copy : alk. paper)
1. Bioinformatics. I. Gopal, Shuba.

QH324.2.B546 2009
572.80285–dc22 2008004080

www.mhhe.com
Dedication
I Mam, Laurel, Sara, Caroline a Claire
—Rhys Price Jones
For Bill, Peter and Scott
—Anne Haake
For Sridhar, Sanjeev and Mara
—Shuba Gopal
Brief Contents

Preface xi
Acknowledgment xiv

Chapter 1 Road Map 1

Chapter 2 Biological Basics 22
Chapter 3 Wet and Dry Lab Techniques 84
Chapter 4 Fragment Assembly 128
Chapter 5 Sequence Alignment 158
Chapter 6 Simulating and Modeling Evolution 219
Chapter 7 Gene Finding 276
Chapter 8 Gene Expression 329
Chapter 9 Projects 383

Index 445

vi
Contents

Preface xi 2.6 Data Maintenance and Integrity

Tasks 70
Acknowledgment xiv 2.6.1 Backing up DNA Data 70
2.6.2 The Challenges of Data
Chapter 1 Road Map 1 Management 74
1.1 What Is Bioinformatics? 1 Key Terms 82
1.2 A Bioinformatics Team 6 Bibliography 83
1.3 What Defines Life? 8
1.4 A Systems Approach to Biology 10 Chapter 3 Wet and Dry Lab
1.5 Bioinformatics in Action 16 Techniques 84
1.5.1 Deciphering a Killer: HIV and
Bioinformatics 17 3.1 Hybridization: Putting Base Pairs to
Work 85
1.6 The Road Ahead 18
3.2 Making Copies of Nucleotide
Summary 20 Sequences 87
Key Terms 20 3.3 An Explosion of Copies 91
Bibliography 21 3.4 Sequencing DNA Strings 94
3.5 The Human Genome Project: Computing to
Chapter 2 Biological Basics 22
the Rescue 99
2.1 The Blind Engineer 22 3.5.1 Mission Impossible: Sequencing the
2.1.1 The Case of the Peppered Human Genome 100
Moth 23 3.6 Human Genome Sequencing
2.1.2 How Evolution Works 24 Strategies 105
2.1.3 Evolution’s Palette 25 3.7 From Structure to Function 107
2.2 Compute Machine par Excellence 26 3.8 Profiling the Transcriptome 111
2.2.1 Cellular Organization and 3.9 A Few Proteomics Techniques 115
Complexity 26 3.10 Putting It All Together 118
2.2.2 Chemistry and Life 28 3.11 A Few Selected Dry Lab Techniques 120
2.2.3 A Parts List for Life 33 3.11.1 Algorithms 120
2.3 The Languages of the Cell 36 3.11.2 Analysis 122
2.3.1 Operating Systems for Cells 37
Key Terms 125
2.3.2 Deciphering the Language of
Bibliography 126
Cells 39
2.3.3 Compiling DNAStrings
Programs 46 Chapter 4 Fragment Assembly 128
2.3.4 Executing Code from 4.1 The Nature of the Problem 128
DNAStrings 49 4.1.1 Two Analogies 128
2.4 Further Nuances in DNAStrings 57 4.1.2 The Need for Multiple
2.5 Proteins: Cellular Machines 61 Coverage 130
2.5.1 Proteins as Molecules 62 4.2 Putting the Pieces Together 132
2.5.2 Proteins as Engineered 4.2.1 Location, Location,
Machines 67 Location 132

vii
viii Contents

4.2.2 Mapping 133 5.4 People Helping Computers:

4.2.3 Using Overlaps 133 Algorithms 176
4.2.4 Whole-Genome Sequencing 134 5.4.1 Alignment 176
4.2.5 The Problem of Repeats 135 5.4.2 Quality of Alignments: Scoring
4.2.6 A Worked Example 135 Schemes 178
4.3 The Size of the Problem 136 5.4.3 Global Alignments: The
Needleman–Wunsch
4.4 A Purely Combinatorial Problem 138
Algorithms 180
4.4.1 Problem Statement 139
5.4.4 A Worked Example 180
4.5 Solving the Combinatorial Problem 139
5.4.5 Local Alignments: The
4.5.1 Overlaps 139
Smith–Waterman Algorithm 187
4.5.2 Fragments Within Fragments 140
5.4.6 A Worked Example 191
4.5.3 A Graph Model 141
5.5 Affine Gap Penalties 192
4.5.4 A Nonoptimal Greedy
Algorithm 142 5.6 Evolutionary Considerations 193
5.6.1 PAM and BLOSUM 193
4.5.5 Improving on Greed 144
5.7 Space/Time Analysis of Dynamic
4.6 Biological Sequence Reassembly 146
Programming Algorithms 196
4.7 Sequencing by Hybridization 148
5.8 Heuristic Approaches: Fast A and
4.7.1 A Worked Example 149
BLAST 197
4.8 Exercises for Chapter 4 155 5.8.1 A Worked Example: Bill Gates
Key Terms 157 at Ballgames 197
Bibliography 157 5.9 Multiple Alignments 199
5.9.1 A Worked Example 202
5.9.2 Analysis of Multiple-Alignment
Chapter 5 Sequence Alignment 158 Algorithms 204
5.10 Exercises for Chapter 5 206
5.1 Exact Pattern Matching 160
5.1.1 The Naïve Algorithm 160 Key Terms 217
5.1.2 Algorithm Analysis 161 Bibliography 218
5.1.3 Other Pattern-Matching
Algorithms 163
5.1.4 DFAs for Search 164
5.1.5 DFAs as Programs 166
Chapter 6 Simulating and Modeling
5.1.6 Suffix Trees 170
Evolution 219
5.1.7 A Worked Example: 6.1 The Biological Time Machine 219
abracadabara 170 6.1.1 Evolutionary Processes 221
5.1.8 Recap of Exact Pattern 6.2 E. coli Evolution 222
Matching 171 6.3 Simulating Evolution in Silico 225
5.2 Things People Do Well: 6.3.1 Genetic Algorithms: A First
Similarity Detection 172 Pass 226
5.3 Computers Helping People: 6.3.2 Monkey Shakespeare: An Extended
Presenting DotPlots 173 Example 228
5.3.1 Straight DotPlot: Searching for 6.3.3 Monkey Evolution: Making Whales
Areas of Exact Matching 173 from Weasels 234
5.3.2 A Worked Example: Can You 6.3.4 A Worked Example: A Genetic
Dance the Can-Can? 174 Algorithm for Gene Finding 238
5.3.3 Controlling Sensitivity and 6.4 Modeling Evolutionary Relationships 241
Selectivity 175 6.4.1 Models of Mutation 243
 Contents ix

6.5 Discovering Evolutionary 8.8 A Worked Example: Gene Expression

Relationships 249 in HIV-Infected Cells 342
6.5.1 Parsimony 252 8.8.1 Data Preprocessing 342
6.5.2 Other Ways to Build Trees 263 8.9 Programs to Work with Genes and
6.5.3 Maximum Likelihood 270 Expression Vectors 350
Key Terms 274 8.10 Mining the Gene Expression Data 352
8.10.1 A Worked Example: Looking for
Bibliography 275
Differentially Expressed
Genes 353
8.10.2 Testing Biological Hypotheses
Chapter 7 Gene Finding 276
with Statistical Hypotheses 354
7.1 A Modern Cryptographic Puzzle 276 8.11 A Worked Example: Forming New
7.1.1 Detecting Encryption 278 Hypotheses 356
7.1.2 Encoding Information 279 8.11.1 Organizing the Data 356
7.2 Cracking the Genome: A First Pass 281 8.11.2 Clustering 358
7.2.1 A Worked Example: HIV 8.11.3 Classification 370
Integration Sites 282 8.11.4 Using Visualization Techniques to
7.2.2 Regulating Genes: Transcription Aid Interpretation 371
Factor-Binding Sites 293 8.11.5 Advanced Classification
7.3 A Biological Decoder Ring 295 Algorithms 373
7.3.1 A First Try at Decryption: ORF 8.12 Data Management 374
Finding 297 8.12.1 Controlled Vocabularies and
7.3.2 Accounting for Discontinuous Standardization of Microarray
Coding Regions 301 Data 375
7.4 Finding Genes Through Mathematics 306 8.13 Exercises for Chapter 8 378
7.4.1 Linguistic Complexity 307
Key Terms 380
7.4.2 Looks Like a . . . 307
Bibliography 381
7.4.3 Markov Models 308
7.4.4 Genes as Markov Processes 314
7.5 Gene Finding by Learning: Letting a
Chapter 9 Projects 383
Computer Do It 317
7.6 Exercises for Chapter 7 319 9.1 Visualization and Exploration of Complex
Datasets 383
Key Terms 327 9.1.1 Sequencing Gel Visualization 384
Bibliography 327 9.1.2 Microarray Data
Visualization 389
9.1.3 Data Visualization Tools 390
Chapter 8 Gene Expression 329 9.1.4 Over to You 392
8.1 Introduction 329 9.1.5 Resources for Visualization 393
8.2 Genes in Context 329 9.2 RNA Structure and Function
8.3 Genotype to Phenotype 330 Prediction 394
8.4 The Expected (by now) Complications of 9.2.1 Solving Structures for Functional
Biology 331 RNAs: Early Successes 395
8.5 A Flood of Data 334 9.2.2 Structural RNAs and Gene
8.6 Noisy Data 337 Regulation 396
8.6.1 Turning down the Noise 338 9.2.3 RNA Structures in Machines:
8.7 The Many Modes of Gene Solving Complex Structures 401
Expression Data 339 9.2.4 Over to You 405
x Contents

9.2.5 Resources for Structure 9.4.3 Over to You 423

Prediction 406 9.4.4 Resources for Information-Based
9.3 Rational Drug Design Through Medicine 424
Protein Structure and 9.5 Systems Biology 424
Function Prediction 408 9.5.1 Introduction 424
9.3.1 A Pharmaceutical Fairy Tale 408 9.5.2 Inputs 428
9.3.2 Drug Development: One in a 9.5.3 Outputs 429
Million Chances 409 9.5.4 Modern Approach to Systems
9.3.3 Structure-Based Drug Biology 429
Design 410 9.5.5 Feedback, Equilibrium, and
9.3.4 A Pharmaceutical Cautionary Attractors 430
Tale 414 9.5.6 What Kind of Model? 435
9.3.5 Over to You 415 9.5.7 Over to You 436
9.3.6 Resources for Rational Drug 9.5.8 Resources for Systems
Design 416 Biology 439
9.4 Information-Based Medicine 416
9.4.1 Identifying Simple Disease Key Terms 440
Genes 417 Bibliography 441
9.4.2 The Challenge of Mapping
Complex Diseases 420 Index 445
Preface

“. . .we seem to agree that [bioinformatics is] an interdisciplinary field, requiring

skills in computer science, molecular biology, statistics, mathematics, and more.
I’m not qualified in any of these fields. . .though I spend most of my time writing
software, developing algorithms, and deriving equations. . .”
— Sean Eddy1

One of the challenges of writing a textbook in an interdisciplinary field as fluid

and rapidly evolving as bioinformatics is that no one is an expert in all aspects
of the field. We all come to the table with different backgrounds, a collection of
skills accrued along the way, and a set of ideas of what to study and how to go
about doing so. To be successful you need to communicate your ideas to people
who share neither the background nor your vocabulary, and to do so in a way
that fires the imagination. That’s what we have tried to do in this book. We are a
disparate group: formal training in mathematics and computer science for two of
us; formal training in biology for the other two; all of us using, developing and
evaluating new computational methods for the analysis of biological problems.
What gets us out of bed in the morning is the idea that we can design an algorithm
or computational method that will help us better understand the miracle of life.
Along the way, we discovered that together we could create a textbook that blends
computing and biology in an engaging and unique way.
In fact, we believe this book is as close to a stand-alone text as possible in the
field. The book is aimed at those with a computing background, but it contains
enough background from both the biological and computing angles that students
from either discipline should feel comfortable starting with this text. Of course,
no text can cover everything in equal depth, so when push comes to shove, we
slide to the computing end of the scale rather than the biology end. Thus, we
assume that students have already acquired about a year’s worth of programming
experience and are conversant with Java or a similar object-oriented programming
language.
To help keep students focused on material that is often complex and detailed,
we used one long-running example throughout the text. Specifically, we pro-
vided worked examples and discussed the ways in which computing approaches
have elucidated key aspects of the biology of the human immunodeficiency virus
(HIV), the virus that causes AIDS. Threading a single example through many top-
ics is meant to enable computing students to ground themselves as they explore
the complexities of biology. In turn, the HIV example gives biology students a
chance to return to a familiar place even as advanced computing concepts are
introduced.

1
Eddy, Sean. (2005). “Ante disciplinary Science,” PLoS Computational Biology. 1(1):e6.

xi
xii Preface

We made a conscious decision not to talk about the application of com-

mon bioinformatics tools, although we highlighted interesting methods that
are embedded in well-known programs. This book is not a “how-to” guide to
bioinformatics, and its intended audience is not computer scientists hoping to
pick up a little biology or biologists hoping to get a few computing clues. Rather,
we hoped to engage the next generation of scientists. We wanted to inspire individ-
uals to design the breakthrough algorithms and compuational approaches that
will provide novel and valuable insights into biological phenomena. Ambitious
as this is, we have learned from our experiences teaching undergraduates that if
you demand the world of them, some will produce amazing results. Indeed, we
wrote this book for just those undergraduates who over the years have surprised
us with their ingenuity, tenacity and enterprising approach to the field. This is a
book aimed at all the undergraduates and possibly first year graduate students
who are just itching for the chance to try out their skills on a whole new set of
problems.
We purposely took a conversational tone in the text, keeping things relatively
lighthearted even as we engaged in rigorous analysis. We hoped in doing so to
entice students into learning and thinking.
We did not think in such lofty terms when we began this process several years
ago. We started at a much more prosaic level: Anne Haake and Rhys Price Jones
were offering a pair of courses titled “Introduction to Bioinformatics Computing”
and “Advanced Bioinformatics Computing” for undergraduates at the Rochester
Institute of Technology (RIT). None of the textbooks on the market had the right
blend of computing and biology for the courses. We identified many textbooks
that were merely procedural guides describing how existing computational tools
could be applied to specific biology problems. Another set of textbooks was so
overwhelmingly focused on the algorithmic aspects that they lost sight of the biol-
ogy that motivated them. We wanted to achieve that happy medium: a textbook
that would emphasize the theoretical underpinnings and logic of the algorithms
while also highlighting the biological knowledge that drives the algorithm devel-
opment in a unique direction. Since no one else was writing the textbook we
wanted, we wrote our own. The fruits of those labors reside in your hands now.
We will not tell you how to use this book because we think there are many
ways to use it. What we do suggest is that you feel free to pick those chapters that
most suit your tastes and the needs of your students. For example, if you or your
students need a little extra biology background, you will find that Chapters 2 and 3
provide a very good primer on basic molecular biology. If, on the other hand,
your students need a gentler introduction to some of the algorithms, chapters 3, 4,
and 5 are designed to help them understand the spirit of bioinformatics algo-
rithms. These chapters do, however, expect a familiarity with written explanations
of algorithmic ideas, such as would be encountered in most introductory
computer science courses at the college level.
You can certainly work through all the chapters in the book over the course
of a semester or year-long course depending on the level of detail you wish to
pursue. You can customize students’ experience by selecting from the exercises
included in each chapter. Many of these exercises require students to develop code
 Preface xiii

and programs to accomplish specific tasks. Most of these tasks have a biological
basis or are motivated by a specific biological phenomenon. For a faster paced,
less involved course, you might omit some of the more detailed programming
exercises. Additional instructional material and solutions to the exercises will be
made available at www.mhhe.com/gopal.
Chapter 9 encourages you to give your students free rein to explore some
fascinating new areas of bioinformatics where computing challenges loom large –
areas in which the extant solutions come in an intriguing variety and the open
questions are just as numerous. Rather than try to cover all the details of ongoing
research, we leave the door open for innovative students to investigate the issues
for themselves, either on their own or in small groups. This provides an important
pedagogical tool: the motivation and context within which students can become
independent investigators in their own right.
In the end, we want this book to be a gateway to new and exciting discoveries.
We invite you to explore, to investigate and then to roam free through the field.
If our experiences are any indication, what you produce from these explorations
will surprise even you. We hope you will enjoy the journey as much as we have.
Shuba Gopal
Rochester, NY, May 2008
Anne Haake
Rochester NY, May 2008
Rhys Price Jones
Washington, D.C., May 2008
Paul Tymann
Rochester, NY, May 2008
 Acknowledgment

“Knowledge is in the end based on acknowledgment.”

—Ludwig Wittgenstein

Many contribute to any endeavor such as this not just those whose names appear
on the cover. These individuals deserve the real credit for bringing this work to
fruition. Our students have been the primary motivation for this work, and we
remain indebted to them. Transforming that inspiration into reality required the
skills of our excellent editorial team at McGraw-Hill. Kelly Lowery and Emily
Lupash helped us get started writing, and we were then ably guided by Alan
Apt. Melinda Bilecki has been a steady source of support through our darkest
hours. Kris Tibbets and Raghu Srinivasan helped us put the final touches on our
manuscript.
We were fortunate to have numerous peer reviewers of our chapters, and
their feedback provided much needed reassurance and valuable suggestions for
improvement. Our student collaborators, Guy Paddock and Eric Foster, pro-
vided some of the code in the text, on the website, and in the Solutions Manual,
while Brendan Dahl helped ensure that exercises were a good fit for the text.
Dr. Robert Parody contributed material and guidance for the statistical analy-
sis of gene expression data. The students of the Introduction to Bioinformatics
Computing and Advanced Bioinformatics Computing courses in Winter 2005
and Spring 2006 reviewed many of the chapters and offered specific suggestions.
Their enthusiasm for this project has buoyed us throughout.
Last but hardly least, we would like to acknowledge the unflagging support
of our near and dear ones. To Laurel, Bill, and Sridhar: you made it easy for us
to dream big.

xiv
 1
Road Map
“Dealing with these system properties, which ultimately must underlie our under-
standing of all cellular behaviour, will require more abstract conceptualisations
than biologists have been used to in the past. We might need to move into a strange
more abstract world…”
—Paul Nurse (Nobel Prize in physiology or medicine, 2001), A long
twentieth century of the cell cycle and beyond, Cell. 2000, 100:71–78.

1.1 WHAT IS BIOINFORMATICS?

Since the thirteenth century scientists have been using the scientific method to for-
mulate a model of the world around us. The scientific method defines a cycle that
starts with an observation of some phenomenon, the development of a hypoth-
esis that describes the phenomenon, and performance of experiments to test the
validity of the hypothesis. Although the hypothesis is the main focus of the scien-
tific method, what really drives scientific advancement is the collection, analysis,
and interpretation of the data generated by the experiments performed to test the
hypothesis.
Gregor Mendel (1822–1884, Figure 1.1), the first person to trace the charac-
teristics of successive generations of pea plants, used the scientific method to test
his theories of heredity. During the middle of his life, Mendel performed hundreds
of experiments on pea plants and observed changes in seven basic characteristics
of the plants, carefully recording the results from each of these experiments in
notebooks. After analyzing the data he collected in his notebooks, Mendel dis-
covered three basic laws1 that govern the passage of a trait from one generation
of a species to another. Although Mendel’s laws are very important in a study of
genetics, what is really important here is the process that Mendel used to develop
these laws, namely, he collected the data produced after performing several experi-
ments, stored the data in his notebooks, and then analyzed the data to develop
his laws.
The basic scientific work being done in laboratories around the world today
follows the same principles that Mendel followed in the 1800s. One thing that has
changed significantly from Mendel’s time is the scale at which we can perform our

1
Mendel’s laws are: the law of dominance, the law of segregation, and the law of independent
assortment.
1
2 Chapter 1 Road Map

FIGURE 1.1
Gregor Mendel.
(© Pixtal / age
Fotostock)

experiments. Mendel’s data were based on the changes that he could observe in the
characteristics of his pea plants from one generation to another. Today biologists
are able to observe the cells and subcellular components of the organisms they
are studying and collect data on the structure of the DNA and other molecules
in these cells. This has not only changed our understanding of some of the basic
processes in living organisms but it has also resulted in a flood of data. Instead
of recording seven characteristics, modern scientists record tens of thousands.
It is no longer possible to use simple notebooks to record the results of these
experiments, let alone analyze the data they produce.
Like scientific experimentation, computers and how they are used have
evolved over time. Charles Babbage (1791–1871, Figure 1.2), a mathematician
and inventor, grew tired of calculating astronomical tables by hand and conceived
of a way to build a mechanical device to perform the calculations automatically.
In 1822, Babbage started work on a computing device—the difference engine
(Figure 1.3)—to automatically calculate mathematical tables. During the course
of his work on the difference engine, he conceived of a more sophisticated
machine he called the analytical engine. The analytical engine was meant to
be programmed using punch cards and would employ features such as sequential
control, branching, and looping. Although Babbage never built a complete work-
ing model of either machine,2 his work became the basis on which many modern
computers are built.

2
One of Babbage’s earlier difference engines was eventually constructed from drawings by a team at
London’s Science Museum in the 1990s. The machine weighs 3 tons and is 10 feet wide by 6 feet tall.
Chapter 1 Road Map 3

FIGURE 1.2
Charles Babbage.
(© Science
Source / Photo
Researchers, Inc.)

FIGURE 1.3
A difference engine.
(© Bettman / Corbis)
4 Chapter 1 Road Map

When electronic computers were first built in the 1940s, they were large, cum-
bersome devices that were only capable of performing simple operations. These
machines were typically very expensive to build and maintain and were designed
for a specific purpose. Like many of the technological advances in the twentieth
century, initially computers were primarily used for military purposes. The face
of computing changed forever with the invention of the transistor, followed by
the integrated circuit. These devices allowed computer engineers to build more
powerful, smaller, and inexpensive machines. These breakthroughs ultimately led
to the development of the personal computer.
In tandem with the development of more powerful, and useful, computer
hardware, computer scientists were also learning more about the basic theories
that underlie computation. Ada Lovelace (1815–1852, Figure 1.4), who also

FIGURE 1.4
Ada Lovelace.
(© Image Select / Art
Resource, NY)
 Chapter 1 Road Map 5

was a mathematician, worked with Charles Babbage on the analytical engine.

Unlike Babbage, who was interested in building a computing device, Lovelace
sought to understand the methodology of computing. She studied these methods,
implementations, and the properties of implementations. Lovelace developed a
program that would have been able to compute the Bernoulli numbers. It is
because of this work that many consider Lovelace to be the world’s first pro-
grammer. It is interesting to note that both Ada Lovelace and Gregor Mendel
were doing their groundbreaking work around the same time.
As computer hardware became more useful and powerful, our understanding
of computing and programming increased as well. New programming languages,
such as Smalltalk, Pascal, and C++, were developed that allow programmers to
write more sophisticated programs. Groundbreaking work at Xerox Palo Alto
Research Center (PARC) produced a computing environment that made it easy
to display information in a graphical form. Computer scientists were developing
an understanding of the basic properties of computation.
The stage was now set for computing and biology to come together. The
curation of the amino acid sequences of proteins begun by Margaret O. Dayhoff
(1925–1983) marked the beginning of modern bioinformatics. Curation involves
the maintenance and annotation of the data, a task well suited to modern
databases. In the Atlas of Protein Sequences, Dayhoff and her colleagues ini-
tiated the formal publication of known sequence data with yearly releases of
the Atlas in the mid-1960s [3]. In 1970, S. B. Needleman and C. D. Wunsch
developed the first algorithm that made it possible to align DNA sequences.
The Needleman–Wunsch algorithm forms the computational basis of sequence
alignment and represents the first major contribution by computer scientists to
the field of bioinformatics. Over the years, computer scientists working together
with biologists have developed faster, more effective algorithms to analyze the
data collected during experimentation.
Computational and quantitative analysis of biological phenomena did not
truly blossom until the explosion of sequence and other biologically derived data
in the mid-1990s. Since then an ocean of biological data has been created. Mod-
ern scientists, unlike Mendel, routinely perform experiments that result in tens of
thousands of data points. For example, using rapid DNA sequencing, scientists
can unravel an entire genome, which typically consists of billions of nucleotides.
A scientist can use a microarray to simultaneously measure the activity levels of
tens of thousands of genes (Figure 1.5). As biotechnology brings ever more effec-
tive techniques to measure biological phenomena, the amount of data produced
by experiments in a laboratory will become even larger, which in turn will require
the development of sophisticated techniques to manage the data, to analyze the
results, and to aid the scientist in interpretation.
Due to the amount and the nature of the data being collected in laboratories
today, biologists are finding more and more that they need to collaborate with
colleagues in different fields such as computer science, information technology,
mathematics, and statistics in order to manage, store, analyze, and visualize the
data produced by experiments they perform in their laboratories. This collabora-
tive work is what makes it possible to interpret the results of modern biological
6 Chapter 1 Road Map

FIGURE 1.5 A microarray. A color version of this image is shown on the inside cover
of this book.
(Courtesy of the Center for Array Technologies at the University of Washington)

experiments and, in turn, develop a better understanding of the nature of living

organisms. This work, done by teams of biologists, chemists, computer scientists,
information technologists, mathematicians, physicists, and statisticians, has given
birth to a new field called bioinformatics.
Bioinformatics is a field of study in which these diverse fields merge into
a single discipline. Working together bioinformaticists develop algorithms and
statistical methods that can be used to analyze and further our understanding
of large collections of different types of biological data, such as nucleotide and
amino acid sequences, protein domains and structures, and the evolutionary
relationship between organisms. During the past decade bioinformaticists have
developed and implemented tools that enable efficient access to and management
of different types of information.
Most of the work in bioinformatics to date has focused on the study of
genomic sequences to provide insight into the function of a cell. Genomic
sequence data is used to predict protein coding regions, amino acid sequences,
and, ultimately, the structure and function of proteins. Although impressive
advances have been made within the past two decades, there is still much that
we do not know or understand. It is clear that advances in the field of bioinfor-
matics rely on a multidisciplinary effort. In the next section we will look at the
challenges faced by those working in this field.

1.2 A BIOINFORMATICS TEAM

Bioinformatics by its very nature requires experts in the various disciplines that
make up this new field. Unfortunately putting together a successful bioinfor-
matics team is not as simple as throwing a few biologists, chemists, computer
scientists, and mathematicians into a room and asking them to work together.
Individuals in these various fields often do not speak the same scientific language
and often only have a basic understanding of the fundamental concepts of the
other disciplines. If the members of a bioinformatics team cannot communicate,
 Chapter 1 Road Map 7

and do not understand or appreciate what the other disciplines offer in terms of
techniques and tools to develop solutions to common problems, the team will
not be successful.
Another way to look at this issue is to consider the spectrum of interests of
the members of the team. In bioinformatics, as in all the sciences, a spectrum
exists that spans the realms of theory, development, and application. The entire
spectrum of both the computational and biological sciences contributes to the
field of bioinformatics, but the boundaries of the realms within the spectrum of
the new science are far from clear. An interesting challenge faces those working
at the interface between the established sciences: If any aspect of the spectrum
of either science is downplayed, then the eventual spectrum of the new discipline
will be deficient.
As an example, consider how the boundaries of theory, development, and
applications come together in a different field—the science of motor mechanics.
Principles of combustion and equations governing energy release in combustive
processes contribute to the realm of the theoretical. How to harness this energy
through an internal combustion or a jet engine falls into the realm of development.
The application realm involves the test pilot or the racing driver. The science of
motor mechanics requires all three aspects of the spectrum. Additionally, any
effective participant in the process—no matter their particular specialty—must
have some knowledge of the rest of the spectrum. The developer needs to know
the theory and the expected eventual use of the product; the test driver needs
an appreciation of the limits of the engine and the processes enabling its power.
Theoreticians must study the consequences and directions of their industry.
For a biologist or a chemist, the creation, study, and analysis of algorithms
are within the theoretical realm. Program design, creation, and testing consti-
tute the developmental realm. Some users of those programs, particularly testers,
participate in the application spectrum of the science of computing. Users who
know and understand the application area will contribute to the modification and
maintenance stages and will need an understanding of the theory and develop-
ment aspects in order to work effectively with the rest of their team. Likewise, for
the computer scientists, information technologists, and statisticians, the chemical
basis of the flow of information within a cell lies in the theoretical realm. How-
ever, a basic understanding of this theory is necessary, to build computational
and statistical models of these processes.
It is important to distinguish the users of motor mechanics—the everyday
drivers and commuters—from the application scientists. Similarly, we distinguish
ordinary users of the computational sciences. Most users of word-processing
programs do not need to know about the underlying representation of their
keystrokes nor about the selection of the algorithms that implement their design
wishes. The term application scientist describes users who contribute to the next
generation of word processors: those who understand the needs of literary com-
position and presentation and who can knowledgeably communicate with the
theoreticians and developers of the products.
Now consider the science of bioinformatics. Where on the spectrum does
the conduct of sequencing experiments belong? What about the design of an
8 Chapter 1 Road Map

efficient database for publishing the results of biological experiments? Is the

search for common features among entries in such a database solely an appli-
cation? Where in the science spectrum of bioinformatics does the design of an
improved alignment program lie?
The truth is that we are as yet unable to specifically identify the boundaries
of the spectrum of this new science. Nor do we need to. What is important is
that all the scientists participating in bioinformatics need to know the entire
potential extent of that spectrum in order to be effective. And the only way
to be certain that you have covered the entire spectrum is to cover the spectra
of both parent sciences: computational science as well as life science. As the
disciplines represented in a bioinformatics team merge, participants in the merged
science whose backgrounds are primarily in one of the parent sciences may be
insufficiently versed in the entire spectrum of the other. And this is a danger
even for those primarily interested in the application aspects of the new sciences.
Application scientists must understand the underlying principles from the realms
of theory and development. Many of the computational applications available
for use today by experimental biologists allow for default assumptions about the
nature of the input and its relevance to the application being used.
For example, the popular basic local alignment search tool (BLAST), used for
sequence similarity searching, is often treated as a “black box” tool that will work
unfailingly regardless of the input and underlying assumptions. To effectively use
BLAST and especially to understand the results it can return, the user needs to
understand principles of evolutionary relatedness and distance as summarized
in the Dayhoff, or point accepted mutation (PAM), matrices (see Chapter 5).
Users who understand the underlying heuristics used to speed up the execution
and the effects of modifying window sizes and thresholds on the usefulness of
the results gain insights into the results returned as well as power, or robustness,
in manipulating the input data correctly. And users with a good understanding
of statistics can have better confidence in their results. Therefore, we emphasize
that bioinformaticists can only benefit from understanding principles across the
entire spectrum of both the biological and computer sciences.
Bioinformatics has come into existence as part of our curiosity to understand
how a biological organism functions. For millennia, humans have been trying to
understand what life is. From Aristotle to Francis Crick and James Watson,
some of the greatest minds have tried to answer this question. Yet, we still do
not have a definitive answer. The next section of this chapter will discuss how an
entire area of science has developed around attempts to define life and identify
its components—the field of biology.

1.3 WHAT DEFINES LIFE?

Because of the nature of the question and the great diversity of life, one aspect
of biology has been largely focused on collecting and describing all the instances
of life available for study. This has led some, Ernest Rutherford in particular, to
dismiss the biological sciences as mere “stamp collecting.” But stamp collecting
 Chapter 1 Road Map 9

can be a valuable activity in this context because it provides the basis for mak-
ing broad generalizations. For example, we can make general statements about
groups of stamps: some have artwork, some are miniaturized photographs, some
are commemorative, and others are decorative, and so on. From these, we might
go on to a theory of stamp “evolution” in which we notice that the cost of stamps
increases over time, or that stamps become more varied in color and tone across
geographical regions. The same approaches are used in biology to draw long-
range inferences about groups of organisms and the changes they undergo across
time and space.
If we ask the question “what is life?” in a different way, we can move past
merely descriptive discussions of what life could be. Let’s ask the question this
way: What defines life?
The answer here is a little bit easier to find. We can say that there are three fea-
tures which are common to all living systems: they are capable of self-reproduction;
they are complex systems; and as a result, they are robust. Self-reproduction is
relatively easy to define; it is the property of being able to create a new copy of
oneself. This, of course, does not mean an exact replica of oneself, and it is, in
fact, the variation between parent and offspring that gives rise to diversity and
drives the changes across time and space known as evolution. This aspect of life is
what best separates it from, say, computers and the common cold virus. Neither
of these can replicate themselves in the absence of direct assistance from a large
cohort of other, possibly living, systems.
It is not difficult to see that most living systems are complex. So, for that mat-
ter, is your desktop or laptop computer. Your computer is made up of millions of
logic circuits and transistors that together create a complex network capable of
sophisticated processing and computation. Similarly, whether you consider a bac-
terium in your stomach or the human brain, each is a complex system capable of
receiving and processing information. This occurs as a result of many small parts
(whether silicon chips or cells) coordinating their responses and communicating
through complex networks to transfer information and responses across time and
space. Unlike computers, which are composed of large numbers of relatively sim-
ple and similar parts, biological systems have large numbers of diverse elements,
many of which can function in more than one way.
Robustness is a more loosely defined concept. When we say that a system
is robust, we mean that it is able to tolerate fluctuations in the environment or
amongst its subcomponents [1]. For example, database management systems are
designed to be robust to the demands of multiple users accessing and writing
information to the database roughly simultaneously. To take a similar example
from biology, human beings are able to maintain body temperatures at about
98.6◦ F regardless of the outside temperature. A high degree of interaction with
the environment is one of the hallmarks that distinguish living systems from most
physical systems. Living organisms require interaction with the environment for
the most basic necessities of life on the one hand and must protect themselves
from harmful environmental influences on the other. Consider the sun, one of
the most important environmental factors. For humans, exposure to sunlight is
essential for vitamin D production yet can cause adverse changes in the skin.
10 Chapter 1 Road Map

Just because a system is robust does not mean that it cannot fail. In our
example, humans can only maintain a normal body temperature within a partic-
ular range of outside temperatures; if it gets too hot or too cold, then the system
fails. Similarly, databases can fail if there is a massive power outage or a catas-
trophic incident that damages the storage system directly. However, in each of
these examples, the system fails only when something goes dramatically wrong.
These systems are able to maintain a stable state most of the time. In other words,
robust systems can account for and manage anticipated perturbation events but
are sensitive to unanticipated and potentially catastrophic failures [2].
Computer systems, such as database management systems, are designed to be
robust. Living systems are continually “reengineered” by evolutionary processes.
In some ways, this is the key definition of living systems: they have evolved to
manage common, “anticipatable” disasters through complex networks designed
to adjust for such fluctuations. Yet the complexity required to manage these
potential disasters leaves them open to rare, albeit dramatic, failures [5, 2]. Take
the case of the dinosaurs. For nearly 300 million years, they dominated the planet.
And then, a single, catastrophic event—the collision of an asteroid—ended their
existence. No matter how robust a system you build, chances are that a single,
extremely low-probability event can completely destroy it. But this leaves the
way open for a new solution to the problem: in the case of the dinosaurs, their
extinction created room for mammals to evolve. Now we humans dominate the
planet. There is no guarantee, however, that we will always remain at the top.
New, more robust organisms might be in the making, just waiting for a chance
to take over as the dominant mode of life.
So how do systems manifest complexity and robustness? The answer is that
many systems are modular in nature. A module is a self-contained unit that can be
modified or evolved, can retain integrity when removed from the larger system,
and that has a clear protocol or method for interfacing with other modules [2, 4].
In the next section we will see how living organisms can be viewed as a collection
of interconnected modules, and how a network of these modules can result in a
complex and robust living organism.

1.4 A SYSTEMS APPROACH TO BIOLOGY

So I went to the librarian in the zoology section and asked her if she could find
me a “map of the cat.” “A map of the cat, sir?” she asked horrified. “You mean a
zoological chart!” From then on there were rumors about a dumb biology student
who was looking for “a map of the cat.”
—Richard P. Feynman, Surely You’re Joking Mr. Feynman!

Living systems utilize networks of interconnected modules to manifest complex-

ity and robustness. As an analogy, you can think of a computer as being one
module in the large network of such modules that are the Internet. Comput-
ers “interact” with one another through the hypertext transfer protocol (http)
or through other, clearly defined, well-ordered protocols. The Internet is an
 Chapter 1 Road Map 11

expansive series of such interconnected modules, and the resulting network is

an information transfer system that is both vast in size and complex in terms of
its connections and responses to changes.
The Internet is a robust system in that it can tolerate perturbations within
the network. Each computer connected to it is a node, and when a node fails,
routers are able to find other ways to transfer the information around that failed
node. In order to disrupt the entire system, a catastrophic event is needed. For
example, a large swath of nodes and routers are suddenly disabled and no re-
routing mechanism can be identified. It is hard to imagine a scenario in which so
many routers and nodes would be disrupted that the entire Internet would fail.
So, by most standards, the Internet is a robust system.
It is even, to some extent, self-replicating in that it grows and expands in
complexity through the addition of new nodes, protocols, and modules. As a
result the Internet is “evolving” across time and space. From an external perspec-
tive, the Internet is constantly changing and adapting to various pressures and
demands, some of which are easy to identify and others more mysterious.
If you were an alien from Mars faced with the challenge of figuring out
how the Internet works, where would you start? You might start by trying to
document all the components of the network and account for the ways in which
each component interacted with other components. Essentially, what you would
be doing is building a “map” of the Internet. This is exactly the approach used
in the Internet mapping project started at Bell Labs in the summer of 1998. The
mapping consists of frequent probes to registered Internet entities. From these
data a tree can be built that shows the paths to most of the nodes on the Internet.
An example of such a map is shown in Figure 1.6.
So, where is the map of the cat or the dog or even of a single cell? It is still a
work in progress. Most of twentieth century biology has been focused on trying to
understand biological phenomena such as development and disease by studying
the behavior of, and more recently by identifying, the underlying molecules. We
have made tremendous progress and have generated huge volumes of data that
remain to be translated into information. Figure 1.7 shows some of the pathways
within an eukaryotic cell such as ours.
A major challenge facing scientists today is to try and assemble the map of
the cellular intra- and internets: the networks that pass information around and
between cells. This is the challenge that you will begin exploring through this text
as we traverse the hierarchy of biological data and examine the computational
approaches to mining the data. The cellular internet is at least as, if not more,
complex than the Internet, and it has had millions of years of tinkering (evolu-
tion) to develop and optimize its connections. The challenge now is essentially to
reverse-engineer this network.
So where are the modules in this biological network? In the context of our
bodies, a cell might be considered as a module because groups of cells create
tissues and organs that result in a human body. But we can zoom into the cell itself,
and it is in here that we find the most astonishing array of modules, networks, and
sophisticated mechanisms for communicating across modules. This is also the
frontier of a new field called systems biology, which today attempts to model
12 Chapter 1 Road Map

FIGURE 1.6 Internet map circa 1998. A more recent version of an internet map is
available at numerous locations, including http://chrisharrison.net/projects/InternetMap/
(© CAIDA / Photo Researchers, Inc.)

the modules within and across cells and to develop a computational replica of
these modules so they can be studied in depth.
What kind of modules would we expect to find in cells? Since self-
reproduction is a crucial aspect of all living organisms, we might expect that
every cell contains a replication module. And indeed, all cells do have such a
module. It is based on the information-carrying molecule in cells, known as DNA
(deoxyribonucleic acid). DNA on its own can’t replicate so there is an associated
set of machines (biologists call these proteins) that are part of a specific network.
The replication module in cells includes DNA, these proteins and all the other
parts required to make a copy of DNA.
We could also think of cells as miniscule computer chips. That is, at every
point, they are receiving input, making decisions as a result of this input, and
generating output (in biology, this is known as responding to a signal, [4]). In cells,
the most common output (response) is to make a machine (protein). Input from
outside the cell is received at the surface of the cell and then transmitted to the
DNA by signaling modules. Thousands of such modules occur in the cell, and all
their signals converge at the DNA. This is where the decisions are made, usually
 Chapter 1 Road Map 13

FIGURE 1.7 Schematic pathways within an eukaryotic cell, as visualized by the VisANT software package.
The software package is described in Hu, Z., Mellor, J., Wu, J. and DeLisi, C. (2005) “VisANT: data-integrating
visual framework for biological networks and modules,” Nucl. Acids Res. 33: W352–W357. The data for this
image was drawn from MouseNet v.1.1 (Kim, Wan K., Krumpelman, Chase, and Marcotte, Edward M. (under
review)). “Inferring mouse gene functions from genomic-scale data using a combined functional network/
classification strategy,” Genome Biology.
(Credit: Courtesy of Zhenjun Hu.)

as a result of combined inputs on the DNA. When a “decision” has been made,
the DNA directs the generation of a short piece of information contained in a
molecule known as RNA (ribonucleic acid), through a process of transcription.
The information in DNA is essentially “transcribed” into another form—the
RNA molecule. The information in the RNA will direct the creation of a protein
through a “protein generation” module in a process known as translation. Rather
than being simply an information-carrying molecule, the resulting protein has its
own unique function and, thus, the information in RNA has been “translated.”
14 Chapter 1 Road Map

The protein can then interact with many other molecules and modules to influence
the behavior of the cell or its neighbors. The output of cells is a change in their
behavior, or the way they respond to the environment. Although today we focus
much of our energy on studying DNA and RNA sequence data, it is the protein
that is the workhorse of the cell and is what “makes us what we are.”
The idea that information flows linearly via information-carrying molecules
is known as the central dogma of molecular biology. It states that information
flows from DNA to RNA to protein. When this idea was originally formulated
in the 1950s, it was thought that information could only flow in one direction:
from DNA to RNA to protein. We’ve since discovered that information can flow
“backwards” as well, from RNA to DNA and from protein to RNA, and we can
take advantage of these alternative flows to learn more about the ways in which
cells receive and respond to their environment. This is one of the best examples of
an important “truth” in biology. Unlike other sciences, biology has very few clear
and unambiguous never-violated principles. The central dogma comes close.
Once a protein is created, the information content inherent in that protein
is transferred when the protein interacts with another protein. These interac-
tions create complex networks. These networks include the signaling modules
that allow information to be received from outside the cell, to be passed around
the cell, and to be transferred to neighboring cells. Given that the web of inter-
actions is so complex and involves thousands or millions of interactions among
diverse, multifunctional elements, the effort to model an entire biological system
is an enormously challenging task. The good news is that, somewhat surprisingly,
molecular biologists have discovered a high degree of unity in these modules, even
among diverse organisms. That is, we can learn a lot about how humans work
by studying fruit flies, worms, or even simpler organisms. Research efforts in
these model organisms, which have fewer cells and well-defined genetic compo-
sitions, contribute greatly to the task of piecing together the puzzle of molecular
interactions.
This also is where computers come in. Although it is difficult for a single
person to track these interactions across time or space, computers can be used to
model what might happen when thousands of proteins interact. In order to model
a biological system, however, we need to know something about each component
of the system as well as how they interact with one another. In Figure 1.8 we
summarize some of the ways in which biologists learn about the components of
a biological module and some of the ways in which computer scientists model
these modules. As you’ll see, the more complex the system gets (e.g., tissues or
organs), the less we know about them from either a biological or computational
standpoint. The future challenges for both computer scientists and biologists lie
in these areas.
What is at stake? Just as computer systems can be engineered to improve their
performance, biologists are attempting to engineer molecular networks, cells, tis-
sues, and even organs in order to improve the quality of life. Take for example,
a cancer cell, which divides uncontrollably and has developed mechanisms to
escape cell death and so keeps on dividing even under adverse environmental
conditions (robustness is not always a good thing). An understanding of the
 Computational
approaches DATA management issues and solutions

Fragment
Neural networks
reassembly
2D & 3D structure
Gene finding
prediction
algorithms
Alignment Phylogeny Graph theory

Machine learning Network modeling

Monte Carlo methods Simulation & modeling Phylogeny
Statistical analyses
DNA RNA Protein Interactions Cells Tissues Organs Systems Organisms

Sequencing 2D & 3D gels Reverse

In situ microscopy Phenotype Symptoms &
Hybridization genetics
observations clinical data
PCR Northern Western blots
blots
Southern blots Mass
spectometry
cDNA synthesis

Biological
approaches

FIGURE 1.8 Computational and biological approaches to modeling organisms.

15
16 Chapter 1 Road Map

modules that control cell division and cell death opens a window for pharmaceu-
tical intervention that will lead scientists to develop new treatments and cures. In
the next section we will take a look at some of the significant projects in bioin-
formatics that have led to a further understanding of how living organisms work
or have improved the quality of life.

1.5 BIOINFORMATICS IN ACTION

The U.S. Human Genome Project (HGP) is one of the best known examples of
bioinformatics in action. The HGP began formally in 1990 and was originally
planned to be a 13-year effort coordinated by the U.S. Department of Energy
and the National Institutes of Health. A genome consists of the DNA in an
organism, including its genes. As you read in Section 1.4, genes carry information
for making most of the proteins required by an organism. One of the goals of the
HGP was to sequence the human genome, which could be used to identify genes
in human DNA. The idea of sequencing the human genome was first proposed
in 1985. At that time portions of the human genome were mapped, which meant
it was possible to determine which portion of the genome was responsible for
some proteins, but no one had a complete list of the sequence of the 3 billion
bases (A, G, C, or T) that made up the genome. Once the human genome was
sequenced, scientists would be able to study the effect of DNA variations among
individuals, which could lead to revolutionary new ways to diagnose, treat, and
someday prevent the thousands of disorders that affect human beings. You may
find it interesting to learn that the Department of Energy played an important
role in the HGP. One of the charges of the DOE is to study the residues of the
atomic bomb project and the biological effects of exposure to radiation. Using
their expertise in engineering technology derived from their massive projects in
atomic energy and its potential dangers, they had already made several major
contributions to the genome project. A previous project initiated by the DOE,
GenBank, was a computerized repository for the primary DNA sequences of
genetic material from all organisms reported in the literature. This was located
at the Los Alamos National Laboratory with a collaborating counterpart at the
European Molecular Biology Laboratories (EMBL) in Heidelberg, Germany.
By 1991, some 60 million bases were recorded, about half of which being human
and the remainder from bacteria, mice, fruit flies, and so forth. This number
doubled over the subsequent 2 years to well over 100 million bases. GenBank
has become the primary repository of sequence data and currently contains over
30 billion bases.
The overwhelming success of the Human Genome Project has resulted in an
explosion in the amount of available biological information, which in turn has
furthered our understanding of the inner working of living organisms. The avail-
ability of this information has dramatically changed the way in which researchers
have been able to do their work. For example, in pharmaceutical research the
traditional pipeline for drug development is a long and expensive one, usually
involving the screening of a large number of potential compounds. Now new
 Chapter 1 Road Map 17

strategies in the process have emerged from our increased understanding of molec-
ular biology and the rapidly growing data on gene sequence, predicted proteins,
and their structures. As opposed to the traditional screening strategies, rational
drug design is a more focused, high-tech, knowledge-based approach. The basic
idea is that if you know exactly which gene and protein is responsible for a dis-
ease, you can custom-make a drug to combat it. Rational drug design uses the
tools of bioinformatics to identify and validate genes and proteins as potential
drug targets. The pharmaceutical industry has embraced genomics as a source of
drug targeting and recognizes that bioinformatics is crucial for validating these
potential drug targets and for determining which ones are the most suitable for
entering the development pipeline. How successful will this be? Time will tell,
but the tremendous potential for success of a rational approach to drug design is
exemplified by the anticancer drugs imatinib mesylate (Gleevec) and trastuzumab
(Herceptin), and the protease inhibitors that are part of the drug cocktail given
to treat individuals infected with HIV. Gleevec, a revolutionary drug, developed
by Novartis Pharmaceuticals and pioneered by Dr. Brian Druker is highly effec-
tive in managing chronic myelogenous leukemia (CML) and has few side effects.
It specifically targets and inhibits a tyrosine kinase, an enzyme that functions
incorrectly in cancer cells. Although the identification of the enzyme inhibitor
and Gleevec’s development relied on more than 10 years of research in the “wet
lab” and clinic, the success of the targeted approach has created a huge amount of
excitement about enzyme inhibitors specifically and about the rational approach,
in general.

1.5.1 Deciphering a Killer: HIV and Bioinformatics

The human immunodeficiency virus (HIV) is the causative agent of AIDS
(acquired immunodeficiency syndrome) and has brought tragedy to millions of
lives. Figuring out the biology of HIV and the secrets of its genome have helped
us develop drugs to fight this deadly agent, but it remains the source of a global
humanitarian and medical crisis. Throughout the book we will examine some
of HIV’s biological complexity and how bioinformatics has helped us decipher
some of its mysteries.
In Chapter 2 we include a description of the biology of HIV. We highlight the
many unusual features of HIV: its RNA genome, its use of reverse transcriptase,
its ability to integrate into the host genome, and its high mutability. Mutability
allows HIV to evade the host immune system. In Chapter 3 we will learn about
some of the laboratory techniques that scientists use to study HIV and its effects
on human cells.
In Chapter 4 we will discuss how retroviruses like HIV are believed to be
the source of many of the repeat regions in the human genome and transposable
elements in many genomes. Although HIV itself has a small genome that was not
difficult to sequence and reassemble, the integration of retroviruses into genomes
bedevils the reassembly process in host genomes.
HIV and its retrovirus compatriots also impinge on the process of sequence
alignment discussed in Chapter 5. Because of their high mutability, different
18 Chapter 1 Road Map

strains of HIV will have slightly different sequences. By subjecting those strains
to multiple alignment, researchers have obtained useful and valuable insights
into where and how HIV originated. The origins of HIV are further explored in
Chapter 6, where we look at methods for determining the evolutionary history
of HIV. We also consider how mutations occur over time in HIV and use this to
build a model of mutation rates.
In Chapter 7, we use HIV as an example of how to identify signals in host
genomes, including our own. HIV integrates itself at specific points within the
host genome, and we will explore the signals that might allow it to select a given
point within the genome as appropriate for integration. This is an example of
how HIV interacts with its host, and these interactions are central to its ability
to infect and spread in a population.
Similarly, in Chapter 8, we consider how HIV influences the gene expression
patterns of a host cell. Because it interferes with host cell protein synthesis, HIV
has a profound effect on gene expression within host cells. Microarray and other
recent technology can track changes in host cell gene expression as a consequence
of HIV infection.
The study of HIV underlies many of the issues raised in the projects detailed
in Chapter 9. In Section 9.2, we consider the secondary structures of RNAs,
including some generated by HIV. These secondary structures are crucial for the
transport of the HIV genome within the cell, and recent work in predicting the
structures of these RNAs has yielded some insights into the biology of HIV.
In Rational Drug Design through Protein Structure and Function Prediction
(Section 9.3), we look at how a computational prediction of the structure of an
HIV protein, the HIV protease, helped to develop a new class of drugs—the
protease inhibitors.
Our hope is that learning about HIV will help you appreciate the many ways
in which bioinformatics can elucidate the mysteries behind a complex biological
system. As we discuss different aspects of HIV biology, you will see that both
computational and experimental methods were used. The study of HIV exempli-
fies the kind of results that can arise when a team of interdisciplinary scientists
bring their skills to bear on a specific problem.

1.6 THE ROAD AHEAD

As we have seen, a successful bioinformatics team is a synergistic combination
of researchers from multiple disciplines who seek insight into the biological pro-
cesses at work in a living organism. Although it is vital that all the members
of a bioinformatics team work together, it is possible to discern two major focus
areas within bioinformatics. The more established of these two areas concentrates
on the biological processes at work within an organism and the use of existing
computational tools to analyze and interpret these data. The second area focuses
on the development of new computing approaches to interpret, manage, and
distribute collections of biological data. The first focus area is biologically moti-
vated and emphasizes collecting information on the state of an organism. The
 Chapter 1 Road Map 19

second studies computing techniques for converting biological data into bio-
logical information. Biologists and chemists are primarily interested in the first
focus area, and computer scientists, information technologists, physicists, and
mathematicians are drawn to the second.
In this book we will follow a “roadmap to systems biology,” taking you on
a journey of discovery from data to information, which is moving us ever closer
to understanding biological problems. You will learn how biological laboratory
science and computational sciences synergistically have enabled a data-driven,
bottom-up approach to extracting information from the data at increasing levels
of complexity. In many cases, computational strategies have greatly acceler-
ated the extraction of information from the data. Just as important, you will
see how effective computational approaches rely on input from the scientific
knowledge base that has been generated from decades of ongoing biological
research.
This book is written for individuals who have already had some training in
the computational sciences. Specifically we assume that you have mastered basic
programming skills and have already encountered basic algorithms and data
structures before starting to read this book. We do not expect that you will have
more than a passing familiarity with biological concepts; these will be developed
in the course of presenting computational concepts. In the chapters that follow we
will introduce both the computational issues, and the biological principals they
rely on, in tandem. The chapters in this book can be divided into three broad
categories: what we term the “kernel” (Chapters 1–3), the “classical” (Chapters 4
and 5), and the “avant-garde” (Chapters 6–9) of bioinformatics.
The first section will cover the requisite background needed by any bioinfor-
maticist. The kernel will develop concepts from molecular biology, methodologies
such as sequencing technology, and other aspects of biology relevant to bio-
informatics. In addition, we will review computing concepts and techniques from
mathematics and statistics. A discussion of database design and use will be part
of this section as well, although databases and their uses, development, and
implementation will be addressed in other chapters as appropriate.
From the kernel you will proceed through the classical bioinformatics section.
This section covers many of the most frequently used bioinformatics tools. We will
focus primarily on the algorithms that define these tools. A secondary emphasis
will be on the biological justifications that motivated the design and development
of the algorithms. Chapters in this section will devote attention to algorithms
involved in fragment reassembly, pattern matching, and dynamic programming.
Many of these methods focus on sequence analysis, an area within bioinformatics
that has seen the greatest pace of development in the past decade.
In the third section of the text, we will extend the analysis to more recent
developments. Some of the topics we cover within the avant-garde bioinformat-
ics section include phylogenetics and tree building (Chapter 6), gene finding
(Chapter 7), clustering of gene expression patterns, and development of gene
networks (Chapter 8). More advanced topics, such as protein structure modeling
and the modeling of cells, tissues, and organs will also be touched on (Chapter 9).
These represent the “future” of bioinformatics and do not yet have formal
20 Chapter 1 Road Map

algorithmic solutions—the area where the most active research will develop in
the years ahead.
When you finish reading this book, you will be conversant with key concepts
in the biological sciences and knowledgeable about current bioinformatics tools
and approaches. Our purpose is not to train you to use those tools. Rather,
you will be able to identify and understand the algorithms that underlie existing
bioinformatics tools and be able to evaluate algorithms and their implementation,
within the context of the relevant biological phenomena.

SUMMARY
Bioinformatics is a new field in which several disci- focus on the biological and chemical processes at
plines work together to manage, store, and analyze work in an organism and concentrate on improv-
the data being produced in laboratories around ing the ability to collect information on the state
the world. Although the process being used to of an organism. Computer scientists, information
advance our understanding of living organisms is technologists, and mathematicians working in a
similar to what Gregor Mendel used in the 1800s, second focus area concentrate on the development
the scale and scope of what is being done today is of computing techniques that convert biological
dramatically different. data into biological information. Working together
For this new discipline to be successful all scientists in both of these focus areas are improv-
members of a bioinformatics team must have a ing the understanding of the biological processes
basic knowledge of the fundamental concepts of at work within an organism.
the disciplines represented by the team and an This text will focus on the computing side of
understanding and appreciation of what each dis- bioinformatics, but it will also introduce basic bio-
cipline brings to the project. We have seen that logical principles as needed. In the next chapter you
not only is it important for the biologist to under- will learn about the central dogma of molecular
stand computer science, it is equally important biology and the basic molecular structure of DNA
for the computer scientist to understand the basic and RNA. You will also obtain a basic knowl-
biological process at work within a living organism. edge of the science of evolution and a general
Although bioinformatics is a relatively new understanding of the concepts related to biologi-
discipline, after four decades two focus areas of the cal information storage, a general overview of gene
discipline have emerged. Biologists and chemists regulation, alleles, SNPs, and genome structure.

KEY TERMS
bioinformatics (1.1) robust (1.3) DNA (deoxyribonucleic acid) (1.4)
theoretical (1.2) evolution (1.3) RNA (ribonucleic acid) (1.4)
development (1.2) complex networks (1.3) transcription (1.4)
application (1.2) robustness (1.3) translation (1.4)
user (1.2) modular (1.3) central dogma of molecular
application scientist (1.2) module (1.3) biology (1.4)
self-reproduction (1.3) systems biology (1.4) genome (1.5)
complex system (1.3) replication module (1.4)
 Chapter 1 Road Map 21

BIBLIOGRAPHY
National Biomedical Research Foundation,
1. J. M. Carlson and John Doyle. Complexity
1978.
and robustness. Proc. Natl. Acad. Sci. USA,
4. H. L. Hartwell, J. J. Hopfield, S. Leibler, and
99:2538–2545, 2002.
A. W. Murray. From molecular to modular
2. Marie E. Csete and John C. Doyle. Reverse
cell biology. Nature, 402(SUPP):C47–C52,
engineering of biological complexity. Science,
1999.
295(5560):1664–1669, 2002.
5. H. Kitano. Systems biology: a brief overview.
3. M. O. Dayhoff. Atlas of Protein Sequence
Science, 295:1662–1664, 2002.
and Structure, vol. 5, supplement 3.
 2
Biological Basics
“The difference [between biology and physics] is one of complexity of design.
Biology is the study of complicated things that give the appearance of having
been designed for a purpose . . . Man-made artefacts like computers and cars . . . are
[also] complicated and obviously designed for a purpose . . . They [can be] treated
as biological objects.”
—Richard Dawkins, The Blind Watchmaker

2.1 THE BLIND ENGINEER

When humans set out to build something, whether it is a skyscraper or a com-
puter, we start by laying out a set of plans. These blueprints have several levels
of detail from the wiring and organization of each constituent part up to the
final appearance of the product. Once the plans are laid out, each part is
designed and built by a team of engineers and specialists. These people are
not necessarily concerned with the final product; what they do best is create
one little part of the larger whole. When that larger whole is finally assembled,
it can appear much greater than the sum of its parts. A good example would
be the supercomputer, Deep Blue, which has taken on mythic qualities since
it beat the reigning world chess champion in a rematch in 1997. (Deep Blue
lost the first confrontation with Gary Kasparov in 1996 and underwent many
upgrades.)
In Chapter 1, we talked about how living systems are defined by this kind
of complexity, in which the whole seems to be much more than the sum of the
parts. In biological systems, the engineers, architects, and design consultants are
replaced by one process: evolution. Whereas human engineers start with a purpose
and design a machine to support that purpose, evolution proceeds with existing
materials with the long-term aim of continued survival. If there is a purpose to
evolutionary processes, then it is simply to propagate life in some form across
time and space.
Success in evolution is defined by the long-term survival of a species or group
of organisms. The important point here is that evolution does not usually operate
on the level of the individual organism, but rather over long periods of time on

22
 Chapter 2 Biological Basics 23

groups of organisms that form a species. Of course, for a species to survive, indi-
vidual organisms are vital. Each individual organism represents a combination
of features that help it survive in a particular environment. These features are
passed from generation to generation across many organisms.
In each generation, some organisms will be better adapted to their environ-
ment, and they will reproduce faster than their less-adapted peers. Over time, the
progeny from the better adapted organisms will begin to dominate the popula-
tion. They will continue to reproduce faster than their less-adapted peers, and
eventually the whole population may exhibit those adaptations that are best for a
particular environment. This is not to say that the population will remain static;
environments do change, and generally those organisms that are flexible enough
to change with their environment will succeed over time.

2.1.1 The Case of the Peppered Moth

A good example of this is a species of moth common in England, Biston betu-
laria. Prior to the Industrial Revolution, the majority of moths had white
wings with small black dots. The peppered moth, as these moths were called,
could hide against the bark of the many light-colored trees that are common
in England. As a result, they were not easily picked out by birds, their main
predators.
Occasionally, a peppered moth might have a variant pattern of entirely black
wings. These moths were easily picked out by birds and often eaten before they
had a chance to reproduce. We would say that the all-black wing feature was a
poor adaptation for the preindustrial environment of England.
Things changed dramatically for these moths when the Industrial Revolution
began spewing coal dust into the air. Now many trees were covered in soot, and
those moths with white wings were distinctive against the blackened bark of trees.
As a result, these moths were easily spotted and picked off by birds. In contrast,
the occasional moth with black wings was perfectly camouflaged and avoided its
predators. Over time, the majority of moths became black-winged because this
was an advantageous adaptation.
Recently, scientists have observed a decline in the numbers of black-winged
moths and a resurgence of white-winged moths. Scientists speculate that the
enforcement of clean air standards in the last half century has once again made
it advantageous to have white wings rather than black wings [9].
So you can see how a feature that is advantageous in certain conditions can
suddenly become deleterious and vice versa.1 Although individual organisms
cannot change their particular set of features, the species as a whole can adapt
to these changes and survive. So when we say that evolution operates on species

1
Recently debate has arisen about this particular example because there is some evidence that the
shift in coloring of the moths cannot be sufficiently explained just by changes in air pollution. For
more on why this might be an oversimplified example, consult Michael Majerus’ book Melanism:
Evolution in Action.
24 Chapter 2 Biological Basics

and not on organisms, we mean that it can drive the adaptations of the species
as whole, but it cannot make a huge difference at the level of the individual
organism.

2.1.2 How Evolution Works

So how does evolution operate? Two processes drive evolution in species: muta-
tion and natural selection. Mutation is the tendency for organisms to change just
a little bit over time. This is the raw material for evolution. In the case of the
peppered moth, a mutation or set of mutations led to the black-wing feature that
was so advantageous during the polluted years prior to clean air laws.
How does evolution use these mutations? Through a process known as natural
selection. This was first described in 1859 by Charles Darwin in his seminal text,
On the Origin of Species. What Darwin noticed is that tiny differences between
individuals can be magnified over time if a particular difference helps one individ-
ual adapt better to an environment than another of its counterparts. We would
say that the black-wing feature of peppered moths was selected for during the
years when pollution was high. The white-wing feature is now being selected for
because of the cleaner air.
You might ask, “Who or what is doing the selecting here?” The answer is
“It depends.” In the case of the peppered moth, the selection comes from the
predators: the birds. The black- or white-wing feature is protective under certain
conditions because it allows the moths to hide from their natural predators. So,
in a sense, the birds are doing the selecting. The same sort of idea applies to many
species: Adaptations that make an organism weaker or less able to adapt will
make these organisms prime targets for predation. So over time the species as a
whole becomes better adapted and better able to avoid its predators. Of course,
the predators are evolving too, so the equilibrium between prey and predators is
constantly shifting.
Juxtaposed with this are the changes to the environment. No environment is
stable over long periods of time. On a geological scale, environments are always
changing: deserts become grasslands that become forests, oceans rise or fall,
mountains are raised up and then eroded. All these changes put pressure on
species: they must adapt to the changes or face extinction. Sometimes, an envi-
ronment suddenly becomes isolated. For example, a species spread out across a
large area might be split in two by floods or volcanic activity. When these sort
of dramatic changes occur, the two populations may begin to evolve separately.
That is, they begin to adapt to their local environments. Eventually, they may
evolve in such different ways that they no longer resemble each other. This is the
process of speciation, or the process of producing new species.
For example, cows and whales actually share an unexpectedly recent common
ancestor. Many millions of years ago, that common ancestor diverged into two
distinct populations: one adapted to the land, the other to the sea. The result
today is two groups of animals that appear to be very different from each other.
Yet, these species share a great deal of the same information and are closely
related biologically.
 Chapter 2 Biological Basics 25

2.1.3 Evolution’s Palette

What do we mean when we say that cows and whales are related? What we
are actually talking about are the sets of heritable information that each organ-
ism within the species has. In the case of cows and whales, they share much of
the same heritable information. In biology, units of heritable information are
called genes. The mutations that evolution uses to select organisms occur within
genes. Because genes are inherited from one generation to the next, the muta-
tions in the genes are also passed across generations. Adaptations are the result
of sets of mutations within genes that allow organisms to survive in their par-
ticular environment. Think of the peppered moths. The coloring of their wings
is driven by specific genes, and mutations in those genes cause the coloring to
change.
Although changes in genes are necessary to make visible changes to an
organism’s appearance, not all changes at the gene level lead to visible changes.
Changes can occur on two levels, and both levels are utilized by evolution.
Small changes in the content of information are achieved by changing the
letter sequences that make up genes. These changes are known as genotypic
changes. Big changes in the appearance of an organism, such as flippers on
a whale as it evolves away from its cow ancestor, are known as phenotypic
changes.
We usually think of evolution as occurring in a slow, steady manner across
millions of years. This is because we look at phenotypic changes, which tend
to take a long time to appear. However, at the genotypic level, changes occur
all the time. Each generation will have slight mutations in its genes, and not
all of these will lead immediately to a change in phenotype. For example,
you and your siblings probably share many phenotypic features such as the
color of your eyes and hair with your parents. Yet, at the genotypic level,
differences between you and your parents and even your siblings also exist.
Some of these were the result of mutations, but many are the result of a
“shuffling” of genetic information at your conception. Every one of your cells
contains information from each of your parents, but some of the parts were
mixed and matched when you were conceived. As a result, you are genetically
unique, even though you share many genetic similarities with your parents. The
combination of mutations and genetic variation in each generation might even-
tually lead to dramatic phenotypic changes, but probably not for another few
millenia.
This is a key feature of evolutionary processes: on the one hand, mutations
accumulate slowly at the genotypic level. On the other hand, the dramatic pheno-
typic changes are seen every now and then. Evolution is what is known in physics
as a stochastic process. That is, small, random changes can be made through a
process such as mutation until a large and dramatic change occurs. At each point,
selection pressure is exerted so that some features are selected for or against. The
easiest examples of selection to understand are phenotypic changes such as the
coloring of peppered moths, but some pressures select for or against certain geno-
typic changes that do not have obvious phenotypic effects. We will return to this
in later chapters.
26 Chapter 2 Biological Basics

2.2 COMPUTE MACHINE PAR EXCELLENCE

“The uniformity of earth’s life, more astonishing than its diversity, is accountable
by the high probability that we derived, originally, from some single cell, fertilized
in a bolt of lightning as the earth cooled.”
— Lewis Thomas, The Lives of a Cell

Leaps of evolution can be seen all across the biological spectrum. One of the most
striking is the apparently sudden switch from single-celled organisms like bacteria
to multicellular ones including humans. We do not fully know why single-celled
organisms suddenly banded together to form multicellular ones. In other words,
we do not yet understand all the pressures that might have favored selection for
multicellular organisms. What we do know is that today, living organisms come
in essentially two flavors: single-celled or multicelled.
The separation of organisms into these two categories also roughly follows
a division of complexity: single-celled organisms are much less complex than
multicelled ones. This does not mean that bacteria are by any means elementary
systems: we still do not understand much of how bacteria function. Figure 2.1
shows a subset of all organisms that we know about today.
Of the three domains of life shown in Figure 2.1, the bacteria and archaea can
be grouped together into a cluster called prokaryotes. The remainder are mem-
bers of a separate cluster called eukaryotes. Traditionally, biologists differentiate
between groups of organisms based on the complexity of their cells.

2.2.1 Cellular Organization and Complexity

The distinction between prokaryotes and eukaryotes lies in how their cells are
organized. Eukaryote cells are divided into lots of little compartments, much

Phylogenetic tree of life

Archaebacteria

Eukaryotes
Eubacteria

Last universal common ancestor

FIGURE 2.1 The tree of life is a representation of how we think organisms are related
to one another. It presupposes a universal common ancestor, a single organism from
which every living species has evolved. That is shown at the center of the tree. Since that
original ancestor, life has evolved into three main domains: eubacteria, archaebacteria,
and eukaryotes.
 Chapter 2 Biological Basics 27

like modern day computers with multiple hardware bays. Each compartment is
called an organelle, or “little organ.” Each organelle has a specific function, and
eukaryotic cells have specialized mechanisms for transporting substances between
their organelles as needed.
Every multicellular organism is made up of eukaryotic cells. All of our
cells are eukaryotic, as are the cells that make up insects, animals, and plants.
Many single-celled eukaryotes are deadly parasites, such as the organism that
causes malaria (Plasmodium falciparum). Many other single-celled eukaryotes
are benign. If you have ever looked at pond scum under a microscope, much of
what you see are single-celled eukaryotes, such as algae, dinoflagellates, Parame-
cium and other species. A sampling of single-celled eukaryotes, also known
as protozoa, is shown in Figure 2.2. You may already know of some single-
celled eukaryotes: yeast, for example, is used in baking and in the making of
beer.
In contrast to eukaryotes, prokaryotes have just one “hardware bay”: all the
contents of the cell are jumbled up in a single compartment. Prokaryotes have
no organelles. Rather, the organization of the prokaryotic cell occurs through
complexes or clusters of substances. Prokaryotes include organisms like the bac-
teria in your gut and the myriad disease-causing bacteria. The archaeabacteria
are also prokaryotes, although many of them have highly unusual biology that is
not seen in any other organisms.
All cells, whether prokaryote or eukaryote, share certain noteworthy features.
You can think of each of these features as a component in a parts list. By the time
you finish this section, you will have a preliminary understanding of the parts
that make up cells and allow them to perform their functions.
Before we can start talking about cells, we need to review some basic chem-
istry specific to living systems. In this section, we will review the nature of atomic
bonds, the basic atoms and molecules that contribute to life, and a little bit about
how cells regulate the interactions between these molecules.

FIGURE 2.2 A
sampling of some
single-celled
eukaryotes.
28 Chapter 2 Biological Basics

The Discovery of Archaeabacteria

The archaebacteria, as they are sometimes known, are believed to be
some of the most ancient organisms on the planet. However, they were dis-
covered only 30 years ago. In 1977, two researchers, Carl Woese and George
Fox, stumbled on a set of organisms that did not easily fall into either the
bacteria or eukaryote divisions. These organisms appeared superficially to
resemble bacteria: they lacked organelles, had circular DNA genomes, and
had many of the same enzymes that bacteria do. However, they also showed
certain features that had until then only been seen in eukaryotes.
The archaeabacteria are primarily characterized by the extreme envi-
ronments in which they are found. Archaeabacteria live around the edges of
sulfur vents many miles below the ocean surface; archaeabacteria “breathe”
methane and die in the presence of oxygen; and they can survive extreme cold
(up to −20◦ C) and heat (over 100◦ C). The more exotic the places researchers
have looked, the more archaeabacteria they have found. One reason we now
think that archaeabacteria may be the most ancient forms of life is that they
seem uniquely adapted for the very harsh conditions that we think must have
existed when life first appeared on this planet [4].
A number of Websites provide additional information about these un-
usual organisms. One excellent place to start is:
http://www.ucmp.berkeley.edu/archaea/archaea.html

2.2.2 Chemistry and Life

All atoms are composed of protons and neutrons, which together form the nucleus
of an atom. Surrounding this nucleus are electrons, which reside in a series
of concentric orbits known as shells. The number of protons and electrons is
exactly equal in all atoms, but different elements have different numbers of pro-
tons and electrons. This observation is encoded in the periodic table of elements
(Figure 2.3).
Each shell of electrons can hold as many as eight electrons, with one excep-
tion: the very first shell can only hold two electrons. Since different types of
atoms have different numbers of electrons, they will have different numbers of
shells. For example, helium (He) has only two electrons, and these are both found
in the first and only electron shell present in He atoms. In contrast, sodium (Na)
has 11 electrons. These are divided as follows: two in the first shell, eight in the
second shell, and one in the third shell (Figure 2.4).
In general, atoms like to have full electron shells as this is energetically favor-
able. In other words, atoms are most stable when they have two electrons in the
first shell and eight in every successive shell. In the case of Na, the extra electron
in the third shell presents a destabilizing force. As a result, Na atoms are willing to
shed the eleventh electron if they can find a willing recipient. One such recipient
might be chlorine (Cl), which has 17 electrons. How would these electrons be
distributed across its shells (Figure 2.5)?
 Chapter 2 Biological Basics 29

1 Atomic number 18
1 Metal 2
1 H 6 He
1.000 2 C Symbol Semimetal 13 14 15 16 17 4.000
12.01
3 4 5 6 7 8 9 10
2 Li Be Nonmetal B C N O F Ne
Atomic weight
6.941 9.012 10.81 12.01 14.01 16.00 19.00 20.18
11 12 13 14 15 16 17 18
3 Na Mg Al Si P S Cl Ar
22.99 24.31 3 4 5 6 7 8 9 10 11 12 26.98 28.09 30.97 32.07 35.45 39.95
19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36
4 K Ca Sc Ti V Cr Mn Fe Co Ni Cu Zn Ga Ge As Se Br Kr
39.10 40.00 44.96 47.55 50.94 52.00 54.94 55.85 58.93 58.69 60.55 65.39 69.72 72.61 74.92 78.96 79.90 83.80
37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54
5 Rb Sr Sy Zr Nb Mo Tc Ru Rh Pd Ag Cd In Sn Sb Te I Xe
85.47 87.62 88.91 91.22 92.91 95.94 98.91 101.1 102.9 106.4 107.9 112.4 114.8 118.7 121.8 127.6 126.9 131.3
55 56 71 72 73 74 75 76 77 78 79 80 81 82 83 84 85 86
6 Cs Ba Lu Hf Ta W Re Os Ir Pt Au Hg Tl Pb Bi Po At Rn
132.9 137.3 175.0 178.5 180.9 183.8 186.2 190.2 192.2 195.1 197.0 200.6 204.4 207.2 209.0 209.0 210.0 222.0
87 88 103 104 105 106 107 108 109 110 111 112 113 114 115 116 117 118
7 Fr Ra Lr Rf Db Sg Bh Hs Mt Uun Uuu Uub Uut Uuq Uup Uuh Uus Uuo
223.0 226.0 262.1 261.1 262.1 263.1 264.1 265.1 268 269 272 277 289 289 293

57 58 59 60 61 62 63 64 65 66 67 68 69 70
6 La Ce Pr Nd Pm Sm Eu Gd Tb Dy Ho Er Tm Yb
138.9 140.1 140.9 144.2 146.9 150.4 152.0 157.3 158.9 162.5 164.9 167.3 168.9 173.0
89 90 91 92 93 94 95 96 97 98 99 100 101 102
7 Ac Th Pa U Np Pu Am Cm Bk Cf Es Fm Md No
227.0 232.0 231.0 236.0 237.0 244.1 245.1 247.1 247.1 251.1 252.0 257.1 258.1 259.1

FIGURE 2.3 The periodic table of elements summarizes some of the features of each element. Elements are
each made up of just one kind of atom, and the properties of these atoms determine the properties of the element.

FIGURE 2.4 This

schematic shows the
11 electrons of
sodium (Na) as they
would appear if we
Na
Atom could see the electron
shells. Each shell is a
concentric orbit
around the nucleus,
which is made up of
protons and neutrons.

FIGURE 2.5 The

chlorine atom has 17
electrons distributed
across three shells.
The lack of a
complete outer shell
makes chlorine atoms
“greedy” and willing
to take electrons from
other atoms.

Chlorine
17 Cl
30 Chapter 2 Biological Basics

When a Na atom and a Cl atom encounter each other, the Na atom gives up
one of its electrons (the outermost one in the third shell) to Cl (which has only
seven in its outermost shell). The result is that now both atoms have a complete
outer shell (Na has eight in its second shell, which is now its outer shell, and Cl
has eight in its third shell, which is its outer shell). However, the Na atom now
has 11 protons with positive charges but retains only 10 electrons. As a result,
it acquires one positive charge and is denoted as a Na+ ion. The Cl atom now
has one extra electron, giving it a negative charge, so it is denoted as a Cl− ion.
Together, the two form an ionic bond, and the formula NaCl indicates the joining
together of these two atoms to create a molecule that is neutral, or does not have
a charge. NaCl, by the way, is the chemical formula for table salt.
In an ionic bond, electrons are fully transferred from one atom to another.
However, in some instances, electrons are shared between atoms rather than fully
transferred. For example, water (H2 O) is made up of two hydrogen atoms and
one oxygen atom. Each hydrogen atom has just one proton and therefore just one
electron. This leaves hydrogen with an unfilled shell (remember that the first shell
takes two electrons). Oxygen, on the other hand, has six protons and electrons.
As a consequence, the two hydrogens and oxygen “share” their electrons. The
electrons in hydrogen spend some of their time in the hydrogen’s shells and some
of their time in the oxygen’s shells. In return, some of oxygen’s electrons also
split their time between the two hydrogens. When electrons are not transferred
outright but shared across a short distance, this is known as a covalent bond.
Figure 2.6 shows how water is formed in a covalent bond.
One of the consequences of sharing electrons between the atoms of water
is that the electrons are sometimes closer to one atom than another. Because

FIGURE 2.6
Water molecules are Covalent bond
formed when two
hydrogens and one 1p
oxygen share their
electrons through
covalent bonds.
Hydrogen

8p
8n Oxygen

Hydrogen

1p

Covalent bond

Bohr Model of H 2O
 Chapter 2 Biological Basics 31

FIGURE 2.7
H
Because of the slight
+
negative charge on
H the oxygen and the
O H slight positive charge
+ on the hydrogens,
H
O water molecules will
H + O assemble into
H structures that create
hydrogen bonds.
These are shaded in
H
O H this image.
H + +
O
H

oxygen is a much larger atom than hydrogen, the electrons do tend to spend more
time on the oxygen side of the molecule. As a result, the oxygen acquires a slight
negative charge. For the same reason, the hydrogens each end up with a slight
positive charge. This is known as a dipole. When several water molecules are
present together, they arrange themselves so that the slightly positive hydrogens
of neighboring molecules are oriented toward the slightly negative oxygens as
shown in Figure 2.7.
The organization of molecules around a dipole creates weak bonds that hold
the molecules in place, but which can be broken quite easily. The weak bonds that
hold water molecules in strings (as shown in Figure 2.7) are known as hydrogen
bonds because they are formed between hydrogens on one water molecule making
weak connections to the oxygen on a different molecule. Hydrogen bonds are most
common in water, but they can occur in any molecule that contains hydrogens
and atoms that can take on a slightly negative charge (like oxygen).

Energy and Chemical Bonds

When ionic or covalent bonds form, it is through a process known as a reaction.
A certain amount of energy is required for the reaction to proceed and the bond
to form. This energy is captured within the bond and released when the bond is
broken. Some bonds require more energy to form than they release when broken,
and these are known as endothermic reactions. In contrast, some bonds require
very little energy to form, but can release large amounts of energy when broken.
These are known as exothermic reactions. For example, the reaction where pure
hydrogen and oxygen combine to form water is extremely exothermic: usually
there is a tremendous release of energy that creates a small explosion.
Of course, for this reaction to occur, the hydrogen and oxygen have to be
heated (i.e., energy is added to the system). The amount of energy required before
a reaction can proceed is known as activation energy. When the activation energy
is very high, as it is in the case of the formation of water from oxygen and
hydrogen, then the reaction does not proceed spontaneously. In contrast, some
reactions have a very low activation energy and can occur nearly spontaneously.
32 Chapter 2 Biological Basics

For example when table salt (NaCl) is dissolved (ionic bonds of NaCl are broken)
in water, little energy is required for the bonds to break.
The strength of a bond is a measure of the activation energy of the reaction
that would break those bonds. The strength of a bond can vary depending on the
environment, however. Take the instance of table salt. The ionic bonds between
Na and Cl seem to be very weak because you can dissolve (break the bonds)
NaCl so quickly in water. However, if you were to try to break NaCl bonds by
heating table salt, you would need to increase the temperature to 800◦ C before
the bonds begin to break. In contrast, covalent bonds will break very easily when
heated. However, in water, the covalent bonds are much stronger than the ionic
bonds. As with many things, the context of a reaction matters as much as the
components of the reaction. In a biological context, almost everything occurs in
a water-based environment. So the strongest bonds we see in biological molecules
are covalent ones. Many ionic bonds occur in this context as well, but they are
weaker because of the water environment present in all living systems.
In living systems, some atoms and molecules are extremely common, whereas
others are rare or never found. There are over 100 known types of atoms (ele-
ments), but only about 25 occur in living systems. The most common elements
found in living systems are hydrogen, carbon, nitrogen, oxygen, and phosphorus.
Together these atoms combine to form molecules. Some of these molecules can
grow in size and complexity to the point where they incorporate many thousands
of atoms. These are known as macromolecules. Some examples of macromolecules
are DNA and large proteins.
Carbon is the most common element in living systems. Indeed, it was orig-
inally thought that any molecule containing carbon had to be associated with
living organisms. Thus, molecules that contain carbon are known as organic.
Although we now know there are ways to create carbon molecules that do not
require living systems, the majority of all carbon compounds are still generated
by living systems.
You will encounter carbon atoms in much of this chapter because they are
at the heart of so many important structures within cells. Given the prevalence
of carbon compounds, chemists and biochemists have developed notations that
allow them to draw and represent them in a sort of shorthand. In the stick figures
in Figure 2.8, the carbons are not shown at all. Rather, each of the vertices of the
hexagonal pattern represents a carbon.

H
H H

H H
H

FIGURE 2.8 Many carbon structures are represented without specifically labeling the
carbon atoms within the structure as shown here for the carbon-based molecule benzene.
Rather, each of the vertices of the hexagon represents a carbon atom that is bound to a
hydrogen (H).
 Chapter 2 Biological Basics 33

Charged molecules

Uncharged
fatty acids

Charged molecules

FIGURE 2.9 The cell membrane of all cells is made up of two layers of fatty acids,
each of which has a charged molecule at its head. The membranes can “roll up” into
small packages, known as micelles. When they enclose small amounts of water or other
contents, they are known as liposomes. When they enclose the entire contents of a cell,
we refer to them as cell membranes.

2.2.3 A Parts List for Life

The Cellular Great Wall
The first feature that all cells share is a barrier that separates the cell’s internal
contents from its environment. In cells, this barrier is known as the cell membrane.
All cells have a cell membrane.2 The cell membrane is actually made up
of two kinds of materials. The internal portion contains fatty acids, which
are hydrophobic, or “water-fearing.” The ends of the fatty acids have charged
molecules, however, that are hydrophilic, or “water-loving.” The charged parti-
cles face outward from the cell and into the cell, and the fatty acid portion faces
into the cell membrane as shown in Figure 2.9. Because there are two layers of
molecules, the cell membrane is known as a bilayer. Because of the hydrophobic
nature of the fatty acid tails, charged particles and large molecules cannot easily
cross the membrane. This circumstance allows the membrane to act as a barrier,
preventing the free movement of molecules.
Why would the cell membrane be so important for the cell’s survival? Cells
are essentially minuscule chemical factories. Every chemical reaction requires just
the right environment in order to occur. The cell membrane creates the first of
several isolated environments that can be used to run various chemical reactions.
Cells are largely made up of water, a charged molecule with some unique
properties. Within this aqueous environment, cells must create and regulate
the flow of energy and information. This is done through a series of chemical
processes that create, maintain, and break bonds between molecules.
Chemical processes usually require specific conditions to be satisfied prior to
the reactions occurring. For example, hydrogen and oxygen, the two components
of water, will not spontaneously form water in any reasonable time.3 For water to

2
Some cells, such as plant cells and some bacteria, have an additional barrier known as the cell wall.
3
It has been calculated that it would take on the order of several billion years for such a spontaneous
reaction to produce even one molecule of water.
34 Chapter 2 Biological Basics

A key feature of the internal environment of the cell is its ability to maintain
a steady state, or equilibrium among hundreds of chemical reactions. In
biology, this is known as homeostasis. Systems that are at homeostasis are
difficult to perturb. That is, you cannot easily shift the equilibrium that the
cell maintains without adding a great deal of energy or force to the system.
For example, your body maintains a steady body temperature around 37◦ C
(98.6◦ F). It takes extreme shifts in the external temperature (about ± 40◦ C)
before your body loses its ability to maintain a steady temperature.

form, the hydrogen and oxygen must be heated to very high temperatures. This
causes an explosive reaction in which water is formed.
In the same manner, the many thousands of chemical reactions required for a
cell to function depend on the creation of the right environment. The temperature
must be correct, the physical and chemical properties of the molecules involved
must be accommodated, and any energy required for the reaction to occur must
be provided. These are just a few of the conditions required for certain reactions to
occur. The cell membrane provides the best mechanism for creating appropriate
environments for chemical reactions to occur within the cell.
A barrier is critical to ensuring that the cell’s contents are protected from
the environment, but the cell needs to know what is going on in its environment.
“Input devices” embedded in the cell membrane communicate changes in the
environment. These input devices, called receptors, are able to “perceive” input
through a chemical interaction: a molecule from the environment chemically
bonds to a portion of the receptor. This initiates changes in the receptor that result
in the transfer of information into the cell. Unlike modern-day computers, cells
do not “read in” data and information from the external environment. Rather,
they usually respond to binary changes in the receptor state. Either the receptor
is “off” and nothing is bound to it, or the receptor is “on” because something is
bound to it.

Cellular Innards
Inside the cell, the various cellular hardware components float in a jelly-like
substance known as the cytoplasm. This is a water-rich environment in which
most of the reactions of the cell are carried out. All the “hardware” of the
cell are embedded in the cytoplasm. The cellular hardware is “welded” to the
cell membrane by special proteins known collectively as the cytoskeleton. These
proteins are able to reorganize their orientation, allowing some cells to move
around their environment. The amoeba, for example, can inch along a sur-
face by extending and contracting parts of its cellular surface, as shown in
Figure 2.10.
In prokaryotic cells, the various hardware is distributed throughout the cell
in a relatively unordered manner. However, in eukaryotic cells, the same hard-
ware is distributed into smaller compartments separated by the same sort of
 Chapter 2 Biological Basics 35

Amoeba extends
part of cellular
1 structure into “pseudopodia”

Movement of cellular contents

New location
of amoeba
3

Distance

FIGURE 2.10 The amoeba moves around in its environment by extending and
contracting parts of the cytoskeleton. As shown in this figure, it begins by extending a
small section of the cell outward in the direction it wishes to go. It then contracts the
“back” of the cell and scoots forward. The amoeba can inch along in this fashion for
extremely long distances, and the entire motion is coordinated by specialized proteins
within the cytoskeleton [1].

membrane that forms the cell membrane. That is, each organelle has its own
membrane.
At the heart of all eukaryotic cells is the nucleus, the main control center and
the place where the full set of cellular instructions known as the genome is stored.
It is also where most of the programs of the cell are first “compiled,” and where
decisions about how to respond to changes in the cell’s environment are made.
We will discuss the genome in much greater detail in the next section.

Energy Sources and Uses

Every cell must have an internal power source or a mechanism for generating
the energy necessary to conduct its daily activities. For most cells, this is done
through a series of chemical reactions in which molecules with high-energy bonds
are created. When these bonds are broken, the energy released can be utilized to
run the various chemical reactions needed for survival, replication, and adap-
tation to the environment. In eukaryotes, the power sources are known as the
mitochondria. Oddly enough, the mitochondria are actually small cells in their
own right. The current theory is that they were prokaryotic microorganisms that
were “swallowed” by a precursor to the eukaryotic cell. The mitochondria now
depend on the larger, eukaryotic cell to provide them with nutrients and other
raw material. In return, they provide the eukaryotic cell with a steady supply of
energy.
Because the mitochondria used to be free-living organisms at some point
in the past, they have their own genomes (sets of programs and instructions for
36 Chapter 2 Biological Basics

NH2

N N
O O O
N N
P P P
HO O O O O
ONa ONa ONa

HO HO

FIGURE 2.11 The energy molecule used in most cells is ATP, adenosine triphosphate.
Each of the three phosphate bonds shown on the left can be broken, releasing 30 kJ of
energy, which can be harnessed to run other chemical reactions within the cell.

running those programs). So every eukaryotic cell has at least two genomes: a
large one stored in the nucleus, and a small one stored in the mitochondria.4
The energy that mitochondria produce is stored in a special molecule known
as adenosine triphosphate (ATP). The energy is actually stored in phosphate bonds
(covalent bonds of three oxygens around a phosphorus atom). This is shown in
Figure 2.11. Each time a phosphate bond is broken, 30 kJ (kilojoules) of energy
is released. Food you consume is eventually converted into these high-energy
bonds in the phosphate groups of ATP. Your cells have a number of complex
biochemical pathways dedicated to the chemical conversion of various food types
into ATP.
Most eukaryotic cells have many more organelles, and each of these has a
specific function. Just as a computer can have various components that provide
added functionality, eukaryotic cells have acquired organelles to handle very
sophisticated tasks. We will discuss some of these organelles as we explore the
workings of the cell. For now, though, we turn to the challenge of getting all these
pieces of hardware to work together. We turn to the “software” needed to run
the cell.

2.3 THE LANGUAGES OF THE CELL

“When a programming language is created that allows programmers to program
in simple English, it will be discovered that programmers cannot speak English.”
—Anonymous
Just as with computers, cells have several “languages” that control the flow
of information. In cells, the equivalent of assembly language is a set of chemical
interactions, which do the “grunt work” of the cell. Although each such inter-
action is crucial for the functioning of the cell and its continued survival, the
language of interaction is rather limited. It is defined by the chemistry of the
components involved and is restricted to those interactions that are physically

4
Plant cells have a third genome stored in a special organelle called the chloroplast, which generates
energy by converting sunlight into molecules with high-energy bonds.
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 In war at least it might have been supposed that the queen would occupy
a subordinate position. Yet in no department of State did Isabella show to
greater advantage than as the organizer of victorious armies; not as a
batallador after the fashion of her distinguished ancestors in Castile and in
Aragon; but as the originator of an entirely new system of military
administration.
Before her time, in Spain, war had been waged by the great nobles and
their retainers in attendance upon the king. There was no such thing as
uniformity of action or preparation, no central organization of any kind.
Each man went into battle to fight and to forage as opportunity offered.
Each commander vied with his fellow nobles in deeds of bravery, and
accorded to them such support as he chose. The sovereign exercised a
general authority, and assumed the active command of the united multitude
of soldiers, on rare and important occasions. If victory followed, as at the
Navas de Tolosa, the soldiers were rewarded with the plunder, and took
possession of the property of the enemy. If the Christians were defeated, the
army melted away; and the king betook himself to the nearest shelter.
But Isabella had no sooner assumed the title of Queen of Castile, than
she was called upon to maintain her pretensions in the field. With no
experience but that of a country palace, with no training but that of a
country cloister, she set herself to work to organize an army. On the 1st of
May, 1474, five hundred horsemen represented the entire forces of the fair
usurper. By the 19th of July she had collected over forty thousand men, had
armed and equipped them ready for the field, and had sent them forward
under the command of Ferdinand to the frontier. Although she was at the
time in delicate health, she was constantly in the saddle, riding long
distances from fortress to fortress, hurrying up recruits all day, dictating
letters all night, giving her zealous personal attention to every detail of
armory and equipment, showing from the first that quiet energy and that
natural aptitude for command that ever so constantly distinguished her. That
her levies were not victorious in no way daunted her determination. A
second army was raised by her, within a few weeks after the first had
melted away under Ferdinand; nor would she listen to any offers of
negotiation, until the enemy had been driven out of Castile.
In the conduct of the war of Granada, with time and money at her
command, her preparations were upon a very different scale. The most
skillful artificers were summoned from every part of Europe to assist in the
work of supplying the army with the necessary material of war. Artillery,
then almost unknown to the military art, was manufactured in Spain
according to the best designs. Model cannon were imported, and the
necessary ammunition collected from abroad. Sword-blades were forged at
home. Not only a commissariat, but a field hospital—institutions till then
unheard of in Spanish warfare—were organized and maintained under the
personal supervision of the queen. The presence of a lady on the day of
battle would, as a rule, as she rightly judged, have been rather a hinderance
than a help; but she was very far from being a mere commissioner of
supply. A first-rate horsewoman, she was constantly seen riding about the
camp, encouraging, inspecting, directing; and in the last days of the siege of
Granada, when the spirits of the troops had begun to flag, she appeared
daily in complete armor, and showed herself upon more than one occasion
in a post of danger on the field. The armies with which Gonsalvo de
Cordova overran Calabria, and annihilated the French at Cerignola, were
prepared and dispatched by Isabella; and if, in a subsequent campaign, the
Great Captain was left without supplies or re-enforcements, it was that the
queen was already sickening to her death, broken down and worn out by her
constant and enormous exertions.
But with all her aptitude for military organization, Isabella had no love
for war. Her first campaign was undertaken to make good her pretensions to
the crown. The extermination of the Moslems was a matter of religious
feeling and patriotic pride, rather than an object of military glory; but she
refused to pursue her conquest across the Straits of Gibraltar. The
expeditions to Italy were a part of Ferdinand’s diplomacy, though the honor
of victory must be shared between Isabella and her Great Captain. But the
queen’s ambition lay not in conquest abroad. On the contrary, as soon as the
last province in Spain had been delivered from the foreign yoke of the
Moor, she turned her attention to the peaceful development of the kingdom;
and, unlettered warrior as she was, she bestowed her royal patronage upon
students and studies, rather than upon the knights and nobles who had
fought her battles before Granada.
The old foundations of the Universities, the new art of printing,
scholarship, music, architecture found in her a generous patron, not so
much from predilection as from policy. Men of letters and men of learning
were welcomed at her court, not only from every part of Spain, but from
every part of Europe. For herself she had little appreciation of literature.
She neither knew nor cared what influence her beloved Inquisition would
have upon science. But as long as the queen lived, learning was honored in
Spain.
In this, as in all other things, her judgment of men was unerring. The
queen who made Gonsalvo the commander-in-chief of her armies, and
Ximenez the president of her council, who selected Torquemada as her
grand inquisitor, and Talavera as her archbishop of Granada, made no
mistake when she invited Peter Martyr to instruct her son in polite letters,
and commissioned Lebrija to compose the first Castilian Grammar for the
use of her court.
Her beauty of face and form are familiar. Yet vanity was unknown to her
nature. Simple and abstemious in her daily life, and despising pomp for its
own sake, no one could make a braver show on fitting occasions; and the
richness of her apparel, the glory of her jewels, and the noble dignity of her
presence, have been celebrated by subjects and strangers.
At the death of Isabella, Ferdinand, in accordance with the provisions of
her will, caused his daughter, Joanna, to be proclaimed queen and himself
regent. Philip, archduke of Austria, the husband of Joanna, having disputed
the rights of his father-in-law and threatened an appeal to arms, the latter in
disgust, with the view of again separating the crowns of Aragon and Castile,
entered into negotiations with Louis XII., married Germaine de Foix, the
niece of Louis (1505), and shortly afterward resigned the regency of
Castile. On the death of Philip, in 1506, he resumed the administration,
though not without opposition, and retained it till his death. In 1508 he
joined the League of Cambray for the partition of Venice, and thus without
any trouble became master of five important Neapolitan cities.
In the following year (1509) the African expedition of Cardinal Ximenez
was undertaken, which resulted in the conquest of Oran. In 1511 Ferdinand
joined Venice and Pope Julius II. in a “holy league” for the expulsion of the
French from Italy. This gave a pretext for invading Navarre, which had
entered into alliance with France, and been laid under Papal interdict in
consequence. Aided by his son-in-law Henry VIII. of England, who sent a
squadron under the Marquis of Dorset to co-operate in the descent on
Guienne, Ferdinand became master of Navarre in 1513; and on June 15,
1515, by a solemn act in Cortes held at Burgos, he incorporated it with the
kingdom of Castile.
 The League of Cambray, which was signed on the 10th of December,
1508, between Louis XII., the Emperor Maximilian, and Ferdinand of
Aragon, at the instance of the warlike Pope Julius II., was nominally
directed against the Turks, but was in reality a coalition for the destruction
and partition among the confiscators of the rich State of Venice. If anything
was wanted to make this league of public plunderers more corrupt and more
odious than it would under any circumstances have been, it was that the
kings of France and of Aragon, in order to secure the adhesion of the
Medicis, sacrificed their faithful allies, the Pisans, after solemn assurances
of protection and support, and actually sold that ancient city to the
Florentines, their hereditary enemies, for a hundred thousand ducats.
But all their bad faith and covetousness was displayed in vain. The
perfidious leaguers could not even trust one another; and the success of the
French arms at Agnadel, in May, 1509, so seriously alarmed both Julius and
Ferdinand that a second treaty was concluded in October, 1511, when the
Pope and the King of Aragon invited the Venetian Republic, for whose
destruction they had leagued themselves together with Louis XII. not three
years before, to assist them in driving the French out of Italy.
Of the consummate skill with which Ferdinand, from the middle of 1509
to the end of 1511, played off his allies and rivals one against the other,
until he had accomplished the central object of his diplomacy in the great
Confederation against Louis XII., we may read in the history of France and
of Italy, of England and of Germany, rather than in the Chronicles of
Aragon. For King Ferdinand pulled the strings that moved the puppets,
while he remained wellnigh hidden himself. But by the end of 1511 the
showman was compelled to make his own appearance upon the stage of
European warfare; and Ferdinand was ever less successful as an actor than
as an impresario. His policy for the past two years had been the formation
of a league against his dearly-beloved uncle-in-law, Louis XII., by the aid
of his dearly-beloved son-in-law, Henry VIII. Queen Katharine, who had
already played the part of embassador to her English father-in-law, was to
make use of her influence over her English husband; and if the queen
should refuse to advise King Henry to go to war with France, her confessor
was to tell her that she was bound as a good Christian to do so.
To coerce the confessor, Ferdinand applied to the Pope; and to control
the Pope, he betrayed to him, in secret, the whole scheme of King Louis
XII. as regarded the plunder of the States of the Church. It is easy to
understand what an effect the communication of the French king’s plans of
spoliation produced upon the excitable and irascible Julius. When he had
learned that he was not only to be robbed of his temporalities, but that he
was to be deposed and imprisoned in case he should prove spiritually
intractable, he hastened, in spite of his age and his infirmities, to traverse
the snow-covered mountains, that he might meet his enemy in the field.
The King of Aragon was a diplomatist who left nothing to chance. He
trusted no man. And if no man trusted him, he never deceived himself by
supposing that any one was simple enough to do so. No detail, however
trifling, was neglected by him in his negotiations. No contingency, however
remote, was left out of sight in his intrigues. And however little we may
respect his character, which was perhaps not much worse than that of some
of his rivals, we cannot refuse to admire his transcendent skill, his infinite
perseverance, his forethought, and his keen appreciation of every shade of
political development. A little honesty would have made him a great man, a
little generosity would have made him a great king. His policy, moreover,
toward the close of his life, is at least worthy of an admiration which has
rarely been extended to it. It was a policy which embraced all Europe in its
scope; and although it had no direct relation to Spain or the Spanish people,
it would be ill to conclude even a brief survey of the history of Spain
without referring to the imperial dreams of the great Spaniard, first of
modern diplomatists, and of his early endeavors to solve more than one of
those questions that still embarrass the foreign policy of modern States: the
establishment of a kingdom of Italy; the alliance between Italy and
Germany, to withstand a dreaded power beyond the Danube and the
Carpathians; the entanglement of England in a central European league; and
the treatment of the Pope of Rome.
The Turks, the medieval bugbear in the East—for the Middle Ages had
also their Eastern Question—were at this time rapidly encroaching upon
Christian Europe; and it was obviously desirable to form a powerful empire,
as a bulwark of Christendom, on the banks of the Danube. The opportunity
of founding a great empire in central Europe actually existed. Ladislaus II.,
king of Bohemia and of Hungary, had only one son, Louis, who was of so
delicate a constitution that no issue could be expected of his marriage. In
case he should die without children, his sister, the Princess Anne, was the
heiress of both his kingdoms; and if her father could be persuaded to marry
her to the heir of the Austrian principalities, Bohemia, Austria, and
Hungary, thus united with the heritage of the Hapsburgs, would form by no
means a contemptible State, which might itself be but the nucleus of a
greater and more ambitious empire.
Naples, which had so lately been added to the Spanish dominions, was
still exposed to the attacks of the French, who claimed one-half, and were
always ready to appropriate to themselves the whole of the kingdom.
Naples was separated from France, indeed, by a considerable extent of
territory in Italy; but the smaller Italian States were too weak to render any
serious resistance, and too fickle to be counted upon as friends or as foes by
any Spanish sovereign. The best way to render Naples secure was, in the
eyes of Ferdinand, the foundation of a great kingdom in northern Italy,
powerful enough to prevent the French from marching their armies to the
south. The formation of such a kingdom moreover would have greatly
facilitated a peaceful division of the great Austro-Spanish inheritance
between Prince Charles and his brother, the Infante Ferdinand.
If Charles could be provided not only with the kingdom of Spain, but
with the possessions of Maximilian and Ladislaus and the Princess Anne,
and the empire of central Europe, his younger brother Ferdinand might
content himself with a kingdom to be made up of all the States of Italy,
protected against the encroachments of France by Spanish infantry and
German landsknechts, and ready to drive the Turk out of the Mediterranean
in support of the Christian empire on the Danube.
The kingdom of Italy, thus designed for his younger grandson by the far-
seeing Ferdinand of Aragon, was to consist of Genoa, Pavia, Milan, and the
Venetian territories on the mainland. The country of the Tyrol, being the
most southern of the Austrian dominions, could, without sensibly
weakening the projected empire, be separated from it and added to the new
kingdom in Italy. Thus stretching from the Mediterranean to the Adriatic,
and from the Gulf of Spezia to the Lake of Constance, this sixteenth century
kingdom of Italy, with the whole power of the Holy Roman Empire to
support it, would have been a splendid endowment for a younger son of the
greatest family on earth. There was also a reasonable prospect that it might
afterward be still further enlarged by the addition of Naples, and the smaller
Italian States would easily have fallen a prey to their powerful neighbor.
But in addition to all this, Ferdinand thought that he would render a notable
service to the Catholic religion and to the peace of Europe if the Church
were thoroughly reformed. What Rome herself has lost by Ferdinand’s
failure it is not given even to the Infallible to know. What the king’s reforms
were to be, we can only shrewdly surmise; and although they would most
assuredly not have been Protestant, they would with equal certainty have
been by no means palatable to the Vatican. For it is reasonably probable that
if either Louis XII. or Ferdinand the Catholic had been permitted to carry
out their designs, the Pope of Rome would have found himself deprived of
his temporal power, and Garibaldi, nay, perchance Luther, would have been
forestalled. It was the reforms of Ximenez that to a large extent prevented
Luther in Spain. The reforms of Ferdinand might possibly have prevented
him in Italy.
It was in 1516 that Ferdinand died. Seven years previous Queen
Germaine had been delivered of a son, who received from his parents the
name of John. But the curse that lay upon the children of Ferdinand was not
yet spent; and the rival of Charles V., the heir of Aragon, Sardinia, Naples,
and Sicily, was permitted to gladden the envious heart of his father by but a
few hours of life. As years passed on there seemed little chance of any
further issue of the King and Queen of Aragon. The unity of Spain at length
appeared to be secure. But the ambition of Ferdinand was even surpassed
by his jealousy. Childless, vindictive, and obstinate, he chafed at the ill-
success of his personal schemes; and rather than suffer the crown of united
Spain to pass over to his daughter’s son and heir, he sought, at the hands of
some medical impostor, the powers that were denied to his old age. The
drug that was to have renewed his youth destroyed his constitution, and his
death was the direct result of one of the least creditable of the many
developments of his jealousy, his obstinacy, and his selfishness.
At length came the inevitable end; and at the wretched hamlet of
Madrigalejo, near Guadalupe, in the mountains of Estremadura, on the 23d
of January of the new year 1516, Ferdinand died; and Spain was at length a
United Kingdom.
 CHAPTER VIII

M O D E R N S PA I N
THE HOUSE OF HAPSBURG—PHILIP II.—DEFEAT OF THE
INVINCIBLE ARMADA—A BOURBON AMONG THEM—
THE PENINSULAR WAR—ALFONSO XIII.
With the death of Ferdinand begins the period of uninterrupted
Hapsburg rule in Spain, which lasted for nearly two centuries. In the course
of this period, the monarchy obtained absolute authority, and Spain, after
rising for a time to be the foremost State in Europe, sank to the position of a
second-rate power, from which it has never since emerged. Aragon and
Castile were distinct kingdoms, and the former was again divided into the
three provinces of Aragon, Catalonia, and Valencia, each of which had its
own Cortes, its own privileges, and the most warmly-cherished traditions of
independence. The foreign possessions of the two crowns were a source of
weakness rather than of strength. France stood ready at the earliest
opportunity to contest the possession of Navarre with Castile, and that of
Naples with Aragon.
The difficulties of domestic government were increased by the fact that
the prospective ruler was a youthful foreigner, who had never visited Spain,
and who was completely ignorant of the customs and even of the language
of the country. Charles—the son of Philip, archduke of Austria, and of Jane,
daughter of Ferdinand and Isabella—had been born and educated in the
Netherlands, of which he had been nominal ruler ever since the death of his
father in 1506. All his friends and advisers were Flemings, who cared
nothing for Spanish interests, and had already acquired an evil reputation
for selfish greed. The first symptom of discontent in Spain was excited by
Charles’s demand to be recognized as king, in utter disregard of his mother.
In Aragon the demand was unhesitatingly refused, but in Castile the
vigorous measures of the famous Cardinal Ximenez secured Charles’s
proclamation.
The regent, however, had great difficulties to face. The nobles, delighted
to be rid of the strong government of Ferdinand, wished to utilize the
opportunity to regain the privileges and independence they had lost. In this
crisis the loyal devotion of Ximenez saved the monarchy. Throwing himself
upon the support of the citizen class, he organized a militia which overawed
the nobles and maintained order. A French invasion of Navarre was
repulsed, and to avoid any danger from the discontent of the inhabitants, all
the fortresses of the province, with the single exception of Pamplona, were
dismantled. These distinguished services were rewarded with more than
royal ingratitude by Charles, who came to Spain in 1517, and who allowed
the aged cardinal to die on November 8th, without even granting him an
interview.
Charles’s enormous inheritance was increased by the successes of Cortes
in Mexico and of Pizarro in Peru, by his own annexation of the Milanese,
and by his conquests in northern Africa.
The glory of Spain was then at its apogee. After his death, which
occurred in 1558, the decline set in. From this time also the House of
Hapsburg became divided into its contemporary branches.
Charles was succeeded by Philip II., his only legitimate son. The
administration of the latter, while successful at home, was a failure abroad.
During his reign a claim to the throne of Portugal was successfully asserted,
and the unity of the Peninsula was completed. Moreover, colonial
possessions were greatly extended. Yet his religious intolerance excited the
revolt of the Netherlands, which resulted in a loss of the seven northern
provinces. His effort to obtain a preponderant influence over France was
dexterously foiled by the succession and triumph of Henry IV. But his great
and historical defeat was that which he experienced with the Armada.
Besides the Spanish crown, Philip succeeded to the kingdoms of Naples
and Sicily, the Duchy of Milan, Franche-Comte, and the Netherlands. In
Africa he possessed Tunis, Oran, the Cape Verd, and the Canary Islands;
and in Asia, the Philippine and Sunda Islands, and a part of the Moluccas.
Beyond the Atlantic he was lord of the most splendid portions of the New
World. The empires of Peru and Mexico, New Spain, and Chili, with their
abundant mines of the precious metals, Hispaniola and Cuba, and many
other of the American islands, were provinces of the sovereign of Spain.
Philip had also the advantage of finding himself at the head of a large
standing army in a perfect state of discipline and equipment, in an age
when, except some few insignificant corps, standing armies were unknown
to Christendom. The renown of the Spanish troops was justly high, and the
infantry in particular was considered the best in the world. His fleet, also,
was far more numerous, and better appointed, than that of any other
European power; and both his soldiers and his sailors had the confidence in
themselves and their commanders which a long career of successful warfare
alone can create.
One nation only had been his active, his persevering, and his successful
foe. England had encouraged his revolted subjects in Flanders against him,
and given them the aid in men and money without which they must soon
have been humbled in the dust. English ships had plundered his colonies;
had defied his supremacy in the New World, as well as the Old; they had
inflicted ignominious defeats on his squadrons; they had captured his cities,
and burned his arsenals on the very coasts of Spain. The English had made
Philip himself the object of personal insult. He was held up to ridicule in
their stage plays and masks, and these scoffs at the man had (as is not
unusual in such cases) excited the anger of the absolute king, even more
vehemently than the injuries inflicted on his power. Personal as well as
political revenge urged him to attack England. Were she once subdued, the
Dutch must submit; France could not cope with him, the empire would not
oppose him; and universal dominion seemed sure to be the result of the
conquest of that malignant island.
For some time the destination of an enormous armament which he had
long been preparing was not publicly announced. Only Philip himself, the
Pope Sixtus, the Duke of Guise, and Philip’s favorite minister, Mendoza, at
first knew its real object. Rumors were sedulously spread that it was
designed to proceed to the Indies to realize vast projects of distant conquest.
Sometimes hints were dropped by Philip’s embassadors in foreign courts
that their master had resolved on a decisive effort to crush his rebels in the
Low Countries. But Elizabeth and her statesmen could not view the
gathering of such a storm without feeling the probability of its bursting on
their own shores. As early as the spring of 1587, Elizabeth sent Sir Francis
Drake to cruise off the Tagus. Drake sailed into the Bay of Cadiz and the
Lisbon Roads, and burned much shipping and military stores, causing
thereby an important delay in the progress of the Spanish preparations.
Drake called this “Singeing the king of Spain’s beard.” Elizabeth also
increased her succors of troops to the Netherlanders, to prevent the Prince
of Parma from overwhelming them, and from thence being at full leisure to
employ his army against her dominions.
Philip had an ally in France who was far more powerful than the French
king. This was the Duke of Guise, the chief of the League, and the idol of
the fanatic partisans of the Romish faith. Philip prevailed on Guise openly
to take up arms against Henry III. (who was reviled by the Leaguers as a
traitor to the true Church, and a secret friend to the Huguenots); and thus
prevent the French king from interfering in favor of Queen Elizabeth. “With
this object, the commander, Juan Iniguez Moreo, was dispatched by him in
the early part of April to the Duke of Guise at Soissons. He met with
complete success. He offered the Duke of Guise, as soon as he took the
field against Henry III., three hundred thousand crowns, six thousand
infantry, and twelve hundred pikemen, on behalf of the king, his master,
who would, in addition, withdraw his embassador from the court of France,
and accredit an envoy to the Catholic party. A treaty was concluded on these
conditions, and the Duke of Guise entered Paris, where he was expected by
the Leaguers, and whence he expelled Henry III. on the 12th of May, by the
insurrection of the barricades. A fortnight after this insurrection, which
reduced Henry III. to impotence, and, to use the language of the Prince of
Parma, did not even ‘permit him to assist the Queen of England with his
tears, as he needed them all to weep over his own misfortunes,’ the Spanish
fleet left the Tagus and sailed toward the British isles.”
Meanwhile in England, from the sovereign on the throne to the peasant
in the cottage, all hearts and hands made ready to meet the imminent deadly
peril. A camp was formed at Tilbury; and there Elizabeth rode through the
ranks, encouraging her captains and her soldiers by her presence and her
words.
The ships of the royal navy at this time amounted to no more than thirty-
six; but the most serviceable merchant vessels were collected from all the
ports of the country; and the citizens of London, Bristol, and the other great
seats of commerce, showed as liberal a zeal in equipping and manning
vessels as the nobility and gentry displayed in mustering forces by land.
The seafaring population of the coast, of every rank and station, was
animated by the same ready spirit; and the whole number of seamen who
came forward to man the English fleet was 17,472. The number of the ships
that were collected was 191; and the total amount of their tonnage 31,985.
There was one ship in the fleet (the “Triumph”) of 1,100 tons, one of 1,000,
one of 900, two of 800 each, three of 600, five of 500, five of 400, six of
300, six of 250, twenty of 200, and the residue of inferior burden.
Application was made to the Dutch for assistance: and, as Stowe expresses
it, “The Hollanders came roundly in, with threescore sail, brave ships of
war, fierce and full of spleen, not so much for England’s aid, as in just
occasion for their own defense; these men foreseeing the greatness of the
danger that might ensue, if the Spaniards should chance to win the day and
get the mastery over them; in due regard whereof their manly courage was
inferior to none.”
The equipment of the Spanish forces consisted of 130 ships (besides
caravels), 3,165 cannon, 8,050 sailors, 2,088 galley-slaves, 18,973 soldiers,
1,382 noblemen, gentlemen, and attendants, 150 monks, with Martin
Alarco, vicar of the Inquisition—the whole under the command of the Duke
of Medina Sidonia.
While this huge armada was making ready in the southern ports of the
Spanish dominions, the Prince of Parma, with almost incredible toil and
skill, collected a squadron of warships at Dunkirk, and his flotilla of other
ships and of flat-bottomed boats for the transport to England of the picked
troops, which were designed to be the main instruments in subduing
England. Thousands of workmen were employed, night and day, in the
construction of these vessels, in the ports of Flanders and Brabant.
One hundred of the kind called hendes, built at Antwerp, Bruges, and
Ghent, and laden with provisions and ammunition, together with sixty flat-
bottomed boats, each capable of carrying thirty horses, were brought, by
means of canals and fosses, dug expressly for the purpose, to Nieuport and
Dunkirk. One hundred smaller vessels were equipped at the former place,
and thirty-two at Dunkirk, provided with twenty thousand empty barrels,
and with materials for making pontoons, for stopping up the harbors, and
raising forts and intrenchments. The army which these vessels were
designed to convey to England amounted to thirty thousand strong, besides
a body of four thousand cavalry, stationed at Courtroi, composed chiefly of
the ablest veterans of Europe; invigorated by rest (the siege of Sluys having
been the only enterprise in which they were employed during the last
campaign), and excited by the hopes of plunder and the expectation of
certain conquest.
 Philip had been advised by the deserter, Sir William Stanley, not to
attack England in the first instance, but first to effect a landing and secure a
strong position in Ireland; his admiral, Santa Cruz, had recommended him
to make sure, in the first instance, of some large harbor on the coast of
Holland or Zealand, where the Armada, having entered the Channel, might
find shelter in case of storm, and whence it could sail without difficulty for
England; but Philip rejected both these counsels, and directed that England
itself should be made the immediate object of attack; and on the 20th of
May the Armada left the Tagus, in the pomp and pride of supposed
invincibility, and amid the shouts of thousands, who believed that England
was already conquered. But steering to the northward, and before it was
clear of the coast of Spain, the Armada was assailed by a violent storm, and
driven back with considerable damage to the ports of Biscay and Galicia. It
had, however, sustained its heaviest loss before it left the Tagus, in the death
of the veteran admiral Santa Cruz, who had been destined to guide it against
England.
This experienced sailor, notwithstanding his diligence and success, had
been unable to keep pace with the impatient ardor of his master. Philip II.
had reproached him with his dilatoriness, and had said with ungrateful
harshness, “You make an ill return for all my kindness to you.” These words
cut the veteran’s heart, and proved fatal to Santa Cruz. Overwhelmed with
fatigue and grief, he sickened and died. Philip II. had replaced him by
Alonzo Perez de Gusman, duke of Medina Sidonia, one of the most
powerful of the Spanish grandees, but wholly unqualified to command such
an expedition. He had, however, as his lieutenants, two seamen of proved
skill and bravery, Juan de Martinez Recalde of Biscay, and Miguel
Orquendo of Guipuzcoa.
On the 12th of July the Armada, having completely refitted, sailed again
for the Channel, and reached it without obstruction or observation by the
English.
The design of the Spaniards was, that the Armada should give them, at
least for a time, the command of the sea, and that it should join the
squadron which Parma had collected off Calais. Then, escorted by an
overpowering naval force, Parma and his army were to embark in their
flotilla, and cross the sea to England, where they were to be landed,
together with the troops which the Armada brought from the ports of Spain.
The scheme was not dissimilar to one formed against England a little more
than two centuries afterward.
The orders of King Philip to the Duke of Medina Sidonia were, that he
should, on entering the Channel, keep near the French coast, and, if
attacked by the English ships, avoid an action, and steer on to Calais Roads,
where the Prince of Parma’s squadron was to join him. The hope of
surprising and destroying the English fleet in Plymouth led the Spanish
admiral to deviate from these orders, and to stand across to the English
shore; but, on finding that Lord Howard was coming out to meet him, he
resumed the original plan, and determined to bend his way steadily toward
Calais and Dunkirk, and to keep merely on the defensive against such
squadrons of the English as might come up with him.
It was on Saturday, the 20th of July, that Lord Effingham came in sight
of his formidable adversaries. The Armada was drawn up in form of a
crescent, which from horn to horn measured some seven miles. There was a
southwest wind; and before it the vast vessels sailed slowly on. The English
let them pass by; and then, following in the rear, commenced an attack on
them. A running fight now took place, in which some of the best ships of
the Spaniards were captured; many more received heavy damage; while the
English vessels, which took care not to close with their huge antagonists,
but availed themselves of their superior celerity in tacking and
maneuvering, suffered little comparative loss.
The Spanish admiral showed great judgment and firmness in following
the line of conduct that had been traced out for him; and on the 27th of July
he brought his fleet unbroken, though sorely distressed, to anchor in Calais
Roads. The Armada lay off Calais, with its largest ships ranged outside,
“like strong castles fearing no assault; the lesser placed in the middle ward.”
The English admiral could not attack them in their position without great
disadvantage, but on the night of the 29th he sent eight fire-ships among
them, with almost equal effect to that of the fire-ships which the Greeks so
often employed against the Turkish fleets in their war of independence. The
Spaniards cut their cables and put to sea in confusion. One of the largest
galeasses ran foul of another vessel and was stranded. The rest of the fleet
was scattered about on the Flemish coast, and when the morning broke, it
was with difficulty and delay that they obeyed their admiral’s signal to
range themselves round him near Gravelines. Now was the golden
opportunity for the English to assail them, and prevent them from ever
letting loose Parma’s flotilla against England; and nobly was that
opportunity used. Drake and Fenner were the first English captains who
attacked the unwieldy leviathans: then came Fenton, Southwell, Burton,
Cross, Raynor, and then the lord-admiral, with Lord Thomas Howard and
Lord Sheffield. The Spaniards only thought of forming and keeping close
together, and were driven by the English past Dunkirk, and far away from
the Prince of Parma, who, in watching their defeat from the coast, must, as
Drake expressed it, have chafed like a bear robbed of her whelps. This was
indeed the last and the decisive battle between the two fleets.
Many of the largest Spanish ships were sunk or captured in the action of
this day. And at length the Spanish admiral, despairing of success, fled
northward with a southerly wind, in the hope of rounding Scotland, and so
returning to Spain without a further encounter with the English fleet. Lord
Effingham left a squadron to continue the blockade of the Prince of Parma’s
armament; but that wise general soon withdrew his troops to more
promising fields of action. Meanwhile the lord-admiral himself and Drake
chased the vincible Armada, as it was now termed, for some distance
northward; and then, when it seemed to bend away from the Scotch coast
toward Norway, it was thought best, in the words of Drake, “to leave them
to those boisterous and uncouth northern seas.”
The sufferings and losses which the unhappy Spaniards sustained in their
flight round Scotland and Ireland are well known. Of their whole Armada
only fifty-three shattered vessels brought back their beaten and wasted
crews to the Spanish coast which they had quitted in such pageantry and
pride.
At the death of Philip, which occurred on September 13, 1598, he left to
his son and successor, Philip III., an empire nominally undiminished, but
unwieldy and internally exhausted. Resources had been squandered. The
attention of the masses had been turned from industry to war. The soldiery
once regarded as invincible had lost their prestige in the Netherland
swamps. Enormous taxes, from which nobles and clergy were exempt, were
multiplied on the people. That being insufficient, Philip III. proved his
orthodoxy by completing the work. In 1609 the Moors, or Moriscoes, as
they were called, were ordered to quit the Peninsula within three days, and
the penalty of death was decreed against all who failed to obey, and against
any Christians who should shelter the recalcitrants.
 The edict was obeyed, but it was the ruin of Spain. The Moriscoes were
the backbone of the industrial population, not only in trade and
manufactures, but also in agriculture. The haughty and indolent Spaniards
had willingly left what they considered degrading employments to their
inferiors. The Moors had introduced into Spain the cultivation of sugar,
cotton, rice and silk. They had established a system of irrigation which had
given fertility to the soil. The province of Valencia in their hands had
become a model of agriculture to the rest of Europe. In manufactures and
commerce they had shown equal superiority to the Christian inhabitants,
and many of the products of Spain were eagerly sought for by other
countries. All these advantages were sacrificed to an insane desire for
religious unity.
The resources of Spain, already exhausted, never recovered from this
terrible blow. Philip III. died in March, 1621. His reign had not been
glorious or advantageous to Spain, but it contrasts favorably with those of
his successors. Spanish literature and art, which had received a great
impulse from the intercourse with foreign countries under previous rulers,
reached their zenith during his lifetime. Three writers have obtained
European fame—Cervantes, who produced the immortal “Don Quixote”
between 1605 and 1613, and two of the most fertile of romantic dramatists,
Lope de Vega and Calderon. In the domain of art, Spain produced two of
the greatest masters of the seventeenth century, Velasquez and Murillo.
Philip II. was succeeded by Philip III. After him came Philip IV. and then
Charles II. Of these monarchs Mignet said: “Philip II. was merely a king.
Philip III. and Philip IV. were not kings, and Charles II. was not even a
man.” The death of the latter precipitated the War of the Succession, the
military operations of which were rendered famous by the military exploits
of Eugene and Marlborough. But this is not the place to recite them. The
chief scenes of hostilities were the Netherlands, Germany and Italy, and
their narration belongs more properly to the histories of these lands. Suffice
it to say that by the Treaty of Utrecht war was concluded in 1711, and Philip
V., a Bourbon, second grandson of Louis XIV., was, in accordance with the
will of Charles II., acknowledged King of Spain. By the same treaty
England gained Gibraltar, while the Spanish Netherlands, Milan, Naples
and Sardinia were ceded to Austria.
With the accession of a Bourbon, Spain entered into a new period of
history, during which it once more played a part in the politics of Europe, as
also in its wars; those, for instance, of the Polish and Austrian successions
—the country meanwhile being additionally embroiled with England.
Philip V. was succeeded by Ferdinand VI., and the latter by Charles III.,
whose death, together with the accession of Charles IV., were contemporary
with the French Revolution. The execution of Louis XVI. made a profound
impression on a country where loyalty was a superstition. Charles IV. was
roused to demand vengeance for the insult to his family. Godoy, the Prime
Minister, could but follow the national impulse; and Spain became a
member of the first coalition against France. But the two campaigns which
ensued provoked the contempt of Europe. They form a catalogue of defeats.
Under the circumstances it is no wonder that Spain followed the example of
Prussia and concluded a treaty of peace.
The next event of importance was Napoleon’s famous coup de main—
the seizure of the Spanish royal family at Bayonne—the jugglery which he
performed with the crown, its transference by him from Ferdinand VII. (son
of Charles IV.) to Joseph Bonaparte, and the revolt of the South American
colonies which that act produced.
Then came the restoration of Spanish independence through England’s
aid; Wellington’s famous campaign; the battles of Ciudad Rodrigo and
Badajos; the entry into Madrid; the retreat of Joseph to Valencia;
Napoleon’s crushing defeat at Leipzig, and Ferdinand’s return from
captivity at Valençay.
The circumstances through which these last-mentioned events were
induced or precipitated, and which are collectively known as the Peninsular
War, originated at the moment when Napoleon was practically master of
Europe. Its whole face was changed. Prussia was occupied by French
troops. Holland was changed into a monarchy by a simple decree of the
French emperor, and its crown bestowed on his brother Louis. Another
brother, Jerome, became King of Westphalia, a new realm built up out of
the electorates of Hesse-Cassel and Hanover. A third brother, Joseph, was
made King of Naples; while the rest of Italy, and even Rome itself, was
annexed to the French empire. It was the hope of effectually crushing the
world-power of Britain which drove him to his worst aggression, the
aggression upon Spain.
Napoleon acted with his usual subtlety. In October, 1807, France and
Spain agreed to divide Portugal between them; and on the advance of their
forces the reigning House of Braganza fled helplessly from Lisbon to a
refuge in Brazil. But the seizure of Portugal was only a prelude to the
seizure of Spain. Charles IV., whom a riot in his capital drove at this
moment to abdication, and his son, Ferdinand VII., were drawn to Bayonne
in May, 1808, and forced to resign their claims to the Spanish crown; while
a French army entered Madrid and proclaimed Napoleon’s brother Joseph
king of Spain.
This high-handed act of aggression was hardly completed when Spain
rose as one man against the stranger; and desperate as the effort of its
people seemed, the news of the rising was welcomed throughout England
with a burst of enthusiastic joy. “Hitherto,” cried Sheridan, a leader of the
Whig opposition, “Bonaparte has contended with princes without dignity,
numbers without ardor, or peoples without patriotism. He has yet to learn
what it is to combat a people who are animated by one spirit against him.”
Tory and Whig alike held that “never had so happy an opportunity existed
in Britain to strike a bold stroke for the rescue of the world”; and Canning
at once resolved to change the system of desultory descents on colonies and
sugar islands for a vigorous warfare in the Peninsula.
The furious and bloody struggle which ensued found its climax at
Vittoria, but it would be difficult to find in the whole history of war a more
thrilling chapter than that which tells of the six great campaigns of which
the war itself was composed.
The Peninsular War was perhaps the least selfish conflict ever waged. It
was not a war of aggrandizement or of conquest. It was fought to deliver
Europe from the despotism of Napoleon. At its close the fleets of Great
Britain rode triumphant, and in the Peninsula between 1808-14 her land
forces fought and won nineteen pitched battles, made or sustained ten fierce
and bloody sieges, took four great fortresses, twice expelled the French
from Portugal and once from Spain. Great Britain expended in these
campaigns more than one hundred million pounds sterling on her own
troops, besides subsidizing the forces of Spain and Portugal. This “nation of
shopkeepers” proved that when kindled to action it could wage war on a
scale and in a fashion that might have moved the wonder of Alexander or of
Cæsar, and from motives too lofty for either Cæsar or Alexander so much as
to comprehend. It is worth while to tell afresh the story of some of the more
picturesque incidents in that great strife.
 On April 6, 1812, Badajos was stormed by Wellington; and the story
forms one of the most tragical and splendid incidents in the military history
of the world. Of “the night of horrors at Badajos,” Napier says, “posterity
can scarcely be expected to credit the tale.” No tale, however, is better
authenticated, or, as an example of what disciplined human valor is capable
of achieving, better deserves to be told. Wellington was preparing for his
great forward movement into Spain, the campaign which led to Salamanca,
the battle in which “forty thousand Frenchmen were beaten in forty
minutes.” As a preliminary he had to capture, under the vigilant eyes of
Soult and Marmont, the two great border fortresses, Ciudad Rodrigo and
Badajos. He had, to use Napier’s phrase, “jumped with both feet” on the
first-named fortress, and captured it in twelve days with a loss of twelve
hundred men and ninety officers.
But Badajos was a still harder task. The city stands on a rocky ridge
which forms the last spur of the Toledo range, and is of extraordinary
strength. The river Rivillas falls almost at right angles into the Guadiana,
and in the angle formed by their junction stands Badajos, oval in shape,
girdled with elaborate defenses, with the Guadiana, five hundred yards
wide, as its defense to the north, the Rivillas serving as a wet ditch to the
west, and no less than five great fortified outposts—Saint Roque,
Christoval, Picurina, Pardaleras, and a fortified bridge-head across the
Guadiana—as the outer zone of its defenses. Twice the English had already
assailed Badajos, but assailed it in vain. It was now held by a garrison five
thousand strong, under a soldier, General Phillipson, with a real genius for
defense, and the utmost art had been employed in adding to its defenses. On
the other hand, Wellington had no means of transport and no battery train,
and had to make all his preparations under the keen-eyed vigilance of the
French. Perhaps the strangest collection of artillery ever employed in a
great siege was that which Wellington collected from every available
quarter and used at Badajos. Of the fifty-two pieces, some dated from the
days of Philip II. and the Spanish Armada, some were cast in the reign of
Philip III., others in that of John IV. of Portugal, who reigned in 1640; there
were 24-pounders of George II.’s day, and Russian naval guns; the bulk of
the extraordinary medley being obsolete brass engines which required from
seven to ten minutes to cool between each discharge.
Wellington, however, was strong in his own warlike genius and in the
quality of the troops he commanded. He employed eighteen thousand men
in the siege, and it may well be doubted whether—if we put the question of
equipment aside—a more perfect fighting instrument than the force under
his orders ever existed. The men were veterans, but the officers on the
whole were young, so there was steadiness in the ranks and fire in the
leading. Hill and Graham covered the siege, Picton and Barnard, Kempt and
Colville led the assaults. The trenches were held by the third, fourth, and
fifth divisions, and by the famous light division. Of the latter it has been
said that the Macedonian phalanx of Alexander the Great, the Tenth Legion
of Cæsar, the famous Spanish infantry of Alva, or the iron soldiers who
followed Cortes to Mexico, did not exceed it in warlike quality.
Wellington’s troops, too, had a personal grudge against Badajos, and had
two defeats to avenge. Perhaps no siege in history, as a matter of fact, ever
witnessed either more furious valor in the assault, or more of cool and
skilled courage in the defense. The siege lasted exactly twenty days, and
cost the besiegers five thousand men, or an average loss of two hundred and
fifty per day. It was waged throughout in stormy weather, with the rivers
steadily rising, and the tempests perpetually blowing; yet the thunder of the
attack never paused for an instant.
Wellington’s engineers attacked the city at the eastern end of the oval,
where the Rivillas served it as a gigantic wet ditch; and the Picurina, a
fortified hill, ringed by a ditch fourteen feet deep, a rampart sixteen feet
high, and a zone of mines, acted as an outwork. Wellington, curiously
enough, believed in night attacks, a sure proof of his faith in the quality of
the men he commanded; and on the eighth night of the siege, at nine
o’clock, five hundred men of the third division were suddenly flung on the
Picurina. The fort broke into a ring of flame, by the light of which the dark
figures of the stormers were seen leaping with fierce hardihood into the
ditch and struggling madly up the ramparts, or tearing furiously at the
palisades. But the defenses were strong, and the assailants fell literally in
scores.
Napier tells how “the axmen of the light division, compassing the fort
like prowling wolves,” discovered the gate at the rear, and so broke into the
fort. The engineer officer who led the attack declares that “the place would
never have been taken had it not been for the coolness of these men” in
absolutely walking round the fort to its rear, discovering the gate, and
hewing it down under a tempest of bullets. The assault lasted an hour, and
in that period, out of the five hundred men who attacked, no less than three
hundred, with nineteen officers, were killed or wounded! Three men out of
every five in the attacking force, that is, were disabled, and yet they won!
There followed twelve days of furious industry, of trenches pushed
tirelessly forward through mud and wet, and of cannonading that only
ceased when the guns grew too hot to be used. Captain MacCarthy, of the
Fiftieth Regiment, has left a curious little monograph on the siege, full of
incidents, half tragic and half amusing, but which show the temper of
Wellington’s troops. Thus he tells how an engineer officer, when marking
out the ground for a breaching-battery very near the wall, which was always
lined with French soldiers in eager search of human targets, “used to
challenge them to prove the perfection of their shooting by lifting up the
skirts of his coat in defiance several times in the course of his survey;
driving in his stakes and measuring his distances with great deliberation,
and concluding by an extra shake of his coat-tails and an ironical bow
before he stepped under shelter!”
On the night of April 6, Wellington determined to assault. No less than
seven attacks were to be delivered. Two of them—on the bridge-head across
the Guadiana and on the Pardaleras—were mere feints. But on the extreme
right Picton with the third division was to cross the Rivillas and escalade
the castle, whose walls rose, time-stained and grim, from eighteen to
twenty-four feet high. Leith with the fifth division was to attack the
opposite or western extremity of the town, the bastion of San Vincente,
where the glacis was mined, the ditch deep, and the scarp thirty feet high.
Against the actual breaches Colville and Andrew Barnard were to lead the
light division and the fourth division, the former attacking the bastion of
Santa Maria and the latter the Trinidad. The hour was fixed for ten o’clock,
and the story of that night attack, as told in Napier’s immortal prose, is one
of the great battle-pictures of literature; and any one who tries to tell the tale
will find himself slipping insensibly into Napier’s cadences.
The night was black; a strange silence lay on rampart and trench, broken
from time to time by the deep voices of the sentinels that proclaimed all
was well in Badajos. “Sentinelle garde à vous,” the cry of the sentinels, was
translated by the British private as “All’s well in Badahoo!” A lighted
carcass thrown from the castle discovered Picton’s men standing in ordered
array, and compelled them to attack at once. MacCarthy, who acted as guide
across the tangle of wet trenches and the narrow bridge that spanned the
Rivillas, has left an amusing account of the scene. At one time Picton
declared MacCarthy was leading them wrong, and, drawing his sword,
swore he would cut him down. The column reached the trench, however, at
the foot of the castle walls, and was instantly overwhelmed with the fire of
the besieged. MacCarthy says we can only picture the scene by “supposing
that all the stars, planets, and meteors of the firmament, with innumerable
moons emitting smaller ones in their course, were descending on the heads
of the besiegers.” MacCarthy himself, a typical and gallant Irishman,
addressed his general with the exultant remark, “ ’Tis a glorious night, sir—
a glorious night!” and, rushing forward to the head of the stormers, shouted,
“Up with the ladders!” The five ladders were raised, the troops swarmed up,
an officer leading, but the first files were at once crushed by cannon fire,
and the ladders slipped into the angle of the abutments. “Dreadful their
fall,” records MacCarthy of the slaughtered stormers, “and appalling their
appearance at daylight.” One ladder remained, and, a private soldier
leading, the eager red-coated crowd swarmed up it. The brave fellow
leading was shot as soon as his head appeared above the parapet; but the
next man to him—again a private—leaped over the parapet, and was
followed quickly by others, and this thin stream of desperate men climbed
singly, and in the teeth of the flashing musketry, up that solitary ladder, and
carried the castle.
In the meanwhile the fourth and light divisions had flung themselves
with cool and silent speed on the breaches. The storming party of each
division leaped into the ditch. It was mined, the fuse was kindled, and the
ditch, crowded with eager soldiery, became in a moment a sort of flaming
crater, and the storming parties, five hundred strong, were in one fierce
explosion dashed to pieces. In the light of that dreadful flame the whole
scene became visible—the black ramparts, crowded with dark figures and
glittering arms, on the one side; on the other, the red columns of the British,
broad and deep, moving steadily forward like a stream of human lava. The
light division stood at the brink of the smoking ditch for an instant, amazed
at the sight. “Then,” says Napier, “with a shout that matched even the sound
of the explosion,” they leaped into it and swarmed up to the breach. The
fourth division came running up and descended with equal fury but the
ditch opposite the Trinidad was filled with water; the head of the division
leaped into it, and, as Napier puts it, “about one hundred of the fusiliers, the
men of Albuera, perished there.” The breaches were impassable. Across the
top of the great slope of broken wall glittered a fringe of sword-blades,
sharp-pointed, keen-edged on both sides, fixed in ponderous beams chained
together and set deep in the ruins. For ten feet in front the ascent was
covered with loose planks, studded with sharp iron points. Behind the
glittering edge of sword-blades stood the solid ranks of the French, each
man supplied with three muskets, and their fire scourged the British ranks
like a tempest.
Hundreds had fallen, hundreds were still falling; but the British clung
doggedly to the lower slopes, and every few minutes an officer would leap
forward with a shout, a swarm of men would instantly follow him, and, like
leaves blown by a whirlwind, they swept up the ascent. But under the
incessant fire of the French, the assailants melted away. One private reached
the sword-blades, and actually thrust his head beneath them till his brains
were beaten out, so desperate was his resolve to get into Badajos. The
breach, as Napier describes it, “yawning and glittering with steel, resembled
the mouth of a huge dragon belching forth smoke and flame.” But for two
hours, and until two thousand men had fallen, the stubborn British persisted
in their attacks. Currie, of the 52d, a cool and most daring soldier, found a
narrow ramp beyond the Santa Maria breach only half-ruined; he forced his
way back through the tumult and carnage to where Wellington stood
watching the scene, obtained an unbroken battalion from the reserve, and
led it toward the broken ramp. But his men were caught in the whirling
madness of the ditch and swallowed up in the tumult. Nicholas, of the
engineers, and Shaw of the 43d, with some fifty soldiers, actually climbed
into the Santa Maria bastion, and from thence tried to force their way into
the breach. Every man was shot down except Shaw, who stood alone on the
bastion. “With inexpressible coolness he looked at his watch, said it was too
late to carry the breaches,” and then leaped down! The British could not
penetrate the breach; but they would not retreat. They could only die where
they stood. The buglers of the reserve were sent to the crest of the glacis to
sound the retreat; the troops in the ditch would not believe the signal to be
genuine, and struck their own buglers who attempted to repeat it.
“Gathering in dark groups, and leaning on their muskets,” says Napier,
“they looked up in sullen desperation at Trinidad, while the enemy, stepping
out on the ramparts, and aiming their shots by the light of fire-balls, which
they threw over, asked as their victims fell, ‘Why they did not come into
Badajos.’ ”
 All this while, curiously enough, Picton was actually in Badajos, and
held the castle securely, but made no attempt to clear the breach. On the
extreme west of the town, however, at the bastion of San Vincente, the fifth
division made an attack as desperate as that which was failing at the
breaches. When the stormers actually reached the bastion, the Portuguese
battalions, who formed part of the attack, dismayed by the tremendous fire
which broke out on them, flung down their ladders and fled. The British,
however, snatched the ladders up, forced the barrier, jumped into the ditch,
and tried to climb the walls. These were thirty feet high, and the ladders
were too short. A mine was sprung in the ditch under the soldiers’ feet;
beams of wood, stones, broken wagons, and live shells were poured upon
their heads from above. Showers of grape from the flank swept the ditch.
The stubborn soldiers, however, discovered a low spot in the rampart,
placed three ladders against it, and climbed with reckless valor. The first
man was pushed up by his comrades; he, in turn, dragged others up, and the
unconquerable British at length broke through and swept the bastion. The
tumult still stormed and raged at the eastern breaches, where the men of the
light and fourth division were dying sullenly, and the men of the fifth
division marched at speed across the town to take the great eastern breach
in the rear. The streets were empty, but the silent houses were bright with
lamps. The men of the fifth pressed on; they captured mules carrying
ammunition to the breaches, and the French, startled by the tramp of the
fast-approaching column, and finding themselves taken in the rear, fled. The
light and fourth divisions broke through the gap hitherto barred by flame
and steel, and Badajos was won!
In that dreadful night assault the English lost three thousand five
hundred men. “Let it be considered,” says Napier, “that this frightful
carnage took place in the space of less than a hundred yards square—that
the slain died not all suddenly, nor by one manner of death—that some
perished by steel, some by shot, some by water; that some were crushed and
mangled by heavy weights, some trampled upon, some dashed to atoms by
the fiery explosions—that for hours this destruction was endured without
shrinking, and the town was won at last. Let these things be considered, and
it must be admitted a British army bears with it an awful power. And false
would it be to say the French were feeble men. The garrison stood and
fought manfully and with good discipline, behaving worthily. Shame there
was none on any side. Yet who shall do justice to the bravery of the British
soldiers or the noble emulation of the officers?... No age, no nation, ever
sent forth braver troops to battle than those who stormed Badajos.”
In addition to Badajos, the siege of Ciudad Rodrigo and of San Sebastian
deserve mention. The annals of strife nowhere record assaults more daring
than those which raged in turn around these three great fortresses. Of them
all that of Badajos was the most picturesque and bloody; that of San
Sebastian the most sullen and exasperating; that of Ciudad Rodrigo the
swiftest and most brilliant. A great siege tests the fighting quality of an
army as nothing else can test it. In the night watches in the trenches, in the
dogged toil of the batteries, and the crowded perils of the breach, all the
frippery and much of the real discipline of an army dissolves. The soldiers
fall back upon what may be called the primitive fighting qualities—the
hardihood of the individual soldier, the daring with which the officers will
lead, the dogged loyalty with which the men will follow. As an illustration
of the warlike qualities in a race by which empire has been achieved,
nothing better can be desired than the story of how the breaches were won
at Ciudad Rodrigo.
At the end of 1811 the English and the French were watching each other
jealously across the Spanish border. The armies of Marmont and of Soult,
sixty-seven thousand strong, lay within touch of each other, barring
Wellington’s entrance into Spain. Wellington, with thirty-five thousand
men, of whom not more than ten thousand men were British, lay within
sight of the Spanish frontier. It was the winter time. Wellington’s army was
wasted by sickness, his horses were dying of mere starvation, his men had
received no pay for three months, and his muleteers none for eight months.
He had no siege train, his regiments were ragged and hungry, and the
French generals confidently reckoned the British army as, for the moment
at least, une quantite negligeable.
And yet at that precise moment, Wellington, subtle and daring, was
meditating a leap upon the great frontier fortress of Ciudad Rodrigo, in the
Spanish province of Salamanca. Its capture would give him a safe base of
operations against Spain; it was the great frontier place d’armes for the
French; the whole siege equipage and stores of the army of Portugal were
contained in it. The problem of how, in the depth of winter, without
materials for a siege, to snatch a place so strong from under the very eyes of
two armies, each stronger than his own, was a problem which might have
taxed the warlike genius of a Cæsar. But Wellington accomplished it with a
combination of subtlety and audacity simply marvelous.
He kept the secret of his design so perfectly that his own engineers never
suspected it, and his adjutant-general, Murray, went home on leave without
dreaming anything was going to happen. Wellington collected artillery
ostensibly for the purpose of arming Almeida, but the guns were
transshipped at sea and brought secretly to the mouth of the Douro. No less
than eight hundred mule-carts were constructed without anybody guessing
their purpose. Wellington, while these preparations were on foot, was
keenly watching Marmont and Soult, till he saw that they were lulled into a
state of mere yawning security, and then, in Napier’s expressive phrase, he
“instantly jumped with both feet upon Ciudad Rodrigo.”
This famous fortress, in shape, roughly resembles a triangle with the
angles truncated. The base, looking to the south, is covered by the Agueda,
a river given to sudden inundations; the fortifications were strong and
formidably armed; as outworks it had to the east the great fortified Convent
of San Francisco, to the west a similar building called Santa Cruz; while
almost parallel with the northern face rose two rocky ridges called the Great
and Small Teson, the nearest within six hundred yards of the city ramparts,
and crowned by a formidable redoubt called Francisco. The siege began on
January 8. The soil was rocky and covered with snow, the nights were
black, the weather bitter. The men lacked intrenching tools. They had to
encamp on the side of the Agueda furthest from the city, and ford that river
every time the trenches were relieved. The 1st, 3d, and light divisions
formed the attacking force; each division held the trenches in turn for
twenty-four hours. Let the reader imagine what degree of hardihood it took
to wade in the gray and bitter winter dawn through a half-frozen river, and,
without fire or warm food, and under a ceaseless rain of shells from the
enemy’s guns, to toil in the frozen trenches, or to keep watch, while the
icicles hung from eyebrow and beard, over the edge of the battery for
twenty-four hours in succession.
Nothing in this great siege is more wonderful than the fierce speed with
which Wellington urged his operations. Massena, who had besieged and
captured the city the year before in the height of summer, spent a month in
bombarding it before he ventured to assault. Wellington broke ground on
January 8, under a tempest of mingled hail and rain; he stormed it on the
night of the 19th.
 He began operations by leaping on the strong work that crowned the
Great Teson the very night the siege began. Two companies from each
regiment of the light division were detailed by the officer of the day,
Colonel Colborne, for the assault. Colborne (afterward Lord Seaton), a cool
and gallant soldier, called his officers together in a group and explained
with great minuteness how they were to attack. He then lanched his men
against the redoubt with a vehemence so swift that, to those who watched
the scene under the light of a wintry moon, the column of redcoats, like the
thrust of a crimson sword-blade, spanned the ditch, shot up the glacis, and
broke through the parapet with a single movement. The accidental
explosion of a French shell burst the gate open, and the remainder of the
attacking party instantly swept through it. There was fierce musketry fire
and a tumult of shouting for a moment or two, but in twenty minutes from
Colborne’s lanching his attack every Frenchman in the redoubt was killed,
wounded, or a prisoner.
The fashion in which the gate was blown open was very curious. A
French sergeant was in the act of throwing a live shell upon the storming
party in the ditch, when he was struck by an English bullet. The lighted
shell fell from his hands within the parapet, was kicked away by the nearest
French in mere self-preservation; it rolled toward the gate, exploded, burst
it open, and instantly the British broke in.
For ten days a desperate artillery duel raged between the besiegers and
the besieged. The parallels were resolutely pushed on in spite of rocky soil,
broken tools, bitter weather, and the incessant pelting of the French guns.
The temper of the British troops is illustrated by an incident which George
Napier—the youngest of the three Napiers—relates. The three brothers
were gallant and remarkable soldiers. Charles Napier in India and elsewhere
made history; William, in his wonderful tale of the Peninsular War, wrote
history; and George, if he had not the literary genius of the one nor the
strategic skill of the other, was a most gallant soldier. “I was a field-officer
of the trenches,” he says, “when a 13-inch shell from the town fell in the
midst of us. I called to the men to lie down flat, and they instantly obeyed
orders, except one of them, an Irishman and an old marine, but a most
worthless drunken dog, who trotted up to the shell, the fuse of which was
still burning, and striking it with his spade, knocked the fuse out; then
taking the immense shell in his hands, brought it to me, saying, ‘There she
is for you, now, yer ’anner. I’ve knocked the life out of the crater.’ ”
 The besieged brought fifty heavy guns to reply to the thirty light pieces
by which they were assailed, and day and night the bellow of eighty pieces
boomed sullenly over the doomed city and echoed faintly back from the
nearer hills, while the walls crashed to the stroke of the bullet. The English
fire made up by fierceness and accuracy for what it lacked in weight; but
the sap made no progress, the guns showed signs of being worn out, and,
although two apparent breaches had been made, the counterscarp was not
destroyed. Yet Wellington determined to attack, and, in his characteristic
fashion, to attack by night. The siege had lasted ten days, and Marmont,
with an army stronger than his own, was lying within four marches. That he
had not appeared already on the scene was wonderful.
In a general order issued on the evening of the 19th Wellington wrote,
“Ciudad Rodrigo must be stormed this evening.” The great breach was a
sloping gap in the wall at its northern angle, about a hundred feet wide. The
French had crowned it with two guns loaded with grape, the slope was
strewn with bombs, hand-grenades and bags of powder; a great mine
pierced it beneath; a deep ditch had been cut between the breach and the
adjoining ramparts, and these were crowded with riflemen. The third
division, under General Mackinnon, was to attack the breach, its forlorn
hope being led by Ensign Mackie, its storming party by General Mackinnon
himself. The lesser breach was a tiny gap, scarcely twenty feet wide, to the
left of the great breach; this was to be attacked by the light division, under
Craufurd, its forlorn hope of twenty-five men being led by Gurwood, and
its storming party by George Napier. General Pack, with a Portuguese
brigade, was to make a sham attack on the eastern face, while a fourth
attack was to be made on the southern front by a company of the 83d and
some Portuguese troops. In the storming party of the 83d were the Earl of
March, afterward Duke of Richmond; Lord Fitzroy Somerset, afterward
Lord Raglan; and the Prince of Orange—all volunteers without
Wellington’s knowledge!
At seven o’clock a curious silence fell suddenly on the battered city and
the engirdling trenches. Not a light gleamed from the frowning parapets, not
a murmur arose from the blackened trenches. Suddenly a shout broke out on
the right of the English attack; it ran, a wave of stormy sound, along the line
of the trenches. The men who were to attack the great breach leaped into the
open. In a moment the space between the hostile lines was covered with the
stormers, and the gloomy, half-seen face of the great fortress broke into a
tempest of fire.
Nothing could be finer than the vehement courage of the assault, unless
it were the cool and steady fortitude of the defense. Swift as was the upward
rush of the stormers, the race of the 5th, 77th, and 94th regiments was
almost swifter. Scorning to wait for the ladders, they leaped into the great
ditch, outpaced even the forlorn hope, and pushed vehemently up the great
breach, while their red ranks were torn by shell and shot. The fire, too, ran
through the tangle of broken stones over which they climbed; the hand-
grenades and powder-bags by which it was strewn exploded. The men were
walking on fire! Yet the attack could not be denied. The Frenchmen—
shooting, stabbing, yelling—were driven behind their intrenchments. There
the fire of the houses commanding the breach came to their help, and they
made a gallant stand. “None would go back on either side, and yet the
British could not get forward, and men and officers falling in heaps choked
up the passage, which from minute to minute was raked with grape from
two guns flanking the top of the breach at the distance of a few yards. Thus
striving, and trampling alike upon the dead and the wounded, these brave
men maintained the combat.”
It was the attack on the smaller breach which really carried Ciudad
Rodrigo; and George Napier, who led it, has left a graphic narrative of the
exciting experiences of that dreadful night. The light division was to attack,
and Craufurd, with whom Napier was a favorite, gave him command of the
storming party. He was to ask for one hundred volunteers from each of the
three British regiments—the 43d, 52d, and the rifle corps—in the division.
Napier halted these regiments just as they had forded the bitterly cold river
on their way to the trenches. “Soldiers,” he said, “I want one hundred men
from each regiment to form the storming party which is to lead the light
division to-night. Those who will go with me come forward!”
Instantly there was a rush forward of the whole division, and Napier had
to take his three hundred men out of a tumult of nearly one thousand five
hundred candidates. He formed them into three companies, under Captains
Ferguson, Jones, and Mitchell. Gurwood, of the 52d, led the forlorn hope,
consisting of twenty-five men and two sergeants. Wellington himself came
to the trench and showed Napier and Colborne, through the gloom of the
early night, the exact position of the breach. A staff-officer, looking on,
said, “Your men are not loaded. Why don’t you make them load?” Napier
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