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 Department
of
Microbiology
Unit no 3
Introduction to

Sequence Alignment databases

Subject name
and code
Bioinformatics
& Biostatistics;
02MB0301
Dr. Purvi M.
Rakhashiya
 Importance of Sequence alignment

• Function or activity of a new gene/protein

• Structure or shape of a new protein
• Location or preferred location of a protein
• Stability of a gene or protein
• Origin of a gene, protein, organelle, organism…
 Similarity and Identity

• The key difference between similarity and identity in

sequence alignment is that similarity is the likeness
(resemblance) between two sequences in comparison
while identity is the number of characters that match
exactly between two different sequences.
 Similarity versus Homology

• Similarity refers to the • Homology refers to shared

likeness or % similarity ancestry
between 2 sequences • Two sequences are
• Similarity of sequences homologous if they are
derived from a common
usually means sharing a
ancestral sequence
statistically measured
• Homology often implies
number of bases or amino
similarity
acids (note that structural, but
• Similarity does not not sequence, similarity
necessarily imply may occur)
homology
 Similarity versus Homology contd..

• Similarity can be quantified

• It is correct to say that two sequences are X%
identical
• It is correct to say that two sequences have a
similarity score of Z
• It is correct to say that two sequences are X%
similar, as long as the criteria for similarity is clear.
 Similarity versus Homology
• Homology cannot be quantified
“Its homologous or it isn’t”
• If two sequences have a high % identity it
is OK to say they are homologous
• It is incorrect to say two sequences have a
homology score of Z
• It is incorrect to say two sequences are X
% homologous or have a homology of X %
 Department
of
Microbiology
Unit no 3
Introduction to

Sequence databases

Alignment-2 Subject name

and code
Bioinformatics
& Biostatistics;
02MB0301
Dr. Purvi M.
Rakhashiya
 Homolog Vs. Paralog Vs. Orthog Vs. Analog

• https://youtu.be/TiKbMw_bKEk
• Convergent Vs. Divergent Evolution
– Divergent evolution occurs when two different species share a
common ancestor but have different characteristics from one
another. Each time one ancestral species diverges into multiple
descendant species it is called speciation. Speciation is an
important result of divergent evolution.
– Convergent evolution is the independent evolution of similar
features in species of different periods or epochs in time.
Convergent evolution creates analogous structures that have
similar form or function but were not present in the last common
ancestor of those groups.
• Homologs
– Sequence homology is the biological homology between DNA,
RNA, or protein sequences, defined in terms of shared ancestry in
the evolutionary history of life.
– Two segments of DNA can have shared ancestry because of three
phenomena: either a speciation event (orthologs), or a duplication
event (paralogs), or else a horizontal (or lateral) gene transfer
event (xenologs).
– Paralogs-Paralogous genes are genes that are related via
duplication events in the last common ancestor (LCA) of the
species being compared. They result from the mutation of
duplicated genes during separate speciation events.
– Orthologs- Homologous sequences are orthologous if they are
inferred to be descended from the same ancestral sequence
separated by a speciation event: when a species diverges into two
separate species, the copies of a single gene in the two resulting
species are said to be orthologous. Orthologs, or orthologous
genes, are genes in different species that originated by vertical
descent from a single gene of the last common ancestor.
 Conservation or variation

• Degree of sequence conservation – reflects the evolutionary relatedness of

different sequences

• Degree of sequence variation - reflects the changes that have occurred

during evolution

• Methods of variations
• Mutation or Substitutions
• Insertions
• Deletions
 From unknown to known

• Sequence alignment provides inference for the relatedness of

two sequences under study

• Identifying the evolutionary relationships between sequences

helps to characterize the function of unknown sequences

• If the two sequences share significant similarity, it is extremely

unlikely that the extensive similarity between the two
sequences has been acquired randomly, meaning that the two
sequences must have derived from a common evolutionary
origin
 Causes for sequence (dis)similarity

mutation: a nucleotide at a certain location is replaced by

another nucleotide (e.g.: ATA → AGA)

insertion: at a certain location one new nucleotide is

inserted in between two existing nucleotides
(e.g.: AA → AGA)

deletion: at a certain location one existing nucleotide

is deleted (e.g.: ACTG → AC-G)

indel: an insertion or a deletion

13
Nucleic Acid Sequence characteristics and
parameters

• DNA has a double helical structure

• The strands are complementary
• Hence there is redundancy in DNA in terms of
information
• But the redundant strand is not same
• The direction of both the strands are different
• Anti-parallel

14
 Complementarity

 The two strands of DNA are reverse complementary

5’ ACGTTACG 3’
3’ TGCAATGC 5’
 Most cellular processes involving DNA occur in the 5’ to 3’ direction

 For every G one strand there is a C on other strand and for every A on one strand
there is a T on other strand

15
 DNA Composition (Rigidity and Flexibility)

• AT content
• GC content

• AT or GC content = x100

• High GC = Rigid DNA

• High AT = Flexible DNA
• High probability to interact with proteins
• ATATACGGGCAGCAGC

16
 Types of Alignment

• Pairwise Sequence Alignment

– Global Alignment
– Local Alignment
• Multiple Sequence Alignment
 Pairwise sequence alignment Multiple sequence alignment

• of sequences compared = 2 • of sequences compared > 2

• Can be global or local • Generally global, but can be local
• Simple algorithm for scoring • Sophisticated algorithm for scoring
– Global – Needleman-Wunsch – Progressive alignment
– Local – Smith-Waterman – Pairwise alignment in loop
– Align two most closely related sequences
– Followed by the next sequence most similar
to the pair and so on
 Pairwise sequence alignment Multiple sequence alignment

• To find conserved regions b/w • To detect regions of variability and

sequences conservation in a family of protein
• Similarity search in a database for • Phylogenetic analysis
homologous sequences • Tools
• Tools – MUSCLE
– BLAST – T-Coffee
– EMBOSS Needle – CLUSTAL-W
– EMBOSS water
 Pairwise sequence alignment

• Goal - To find the best pairing of two sequences, such that there is
maximum correspondence among residues

• How - One sequence is shifted relative to the other to find the position
where maximum matches are found

• Strategies -
1. Global alignment
2. Local alignment
 Global Alignment Local Alignment
Local alignments are more useful for
Attempt to align every residue in every dissimilar sequences that are suspected to
sequence, are most useful when the contain regions of similarity or similar
sequences in the query set are similar and of sequence motifs within their larger
roughly equal size. sequence context.
• Assumption – Similarity over the entire • Assumption – Similarity, confined to local
length of sequences regions instead of entire length of
• Sequences – More or less of similar sequences
length • Sequences – could vary largely over
• Alignment - Carried out from beginning length
to end of both sequences • Alignment – carried out considering local
• Aim - Look for best possible alignment regions of high similarity without regard
across the entire length between the two for the alignment of the rest of the
sequences sequence regions
• General global alignment technique is • Aim – Look for local regions with the
the Needleman–Wunsch algorithm highest level of similarity
• General global alignment technique is
the Smith–Waterman algorithm
 Global Alignment Local Alignment

• Applications - More applicable for aligning • Applications - Used for aligning more
two closely related sequences of roughly divergent sequences to find conserved
the same length patterns also known as domains or
• Limitation – Not good for divergent motifs
sequences and sequences of variable
lengths

“:” indicates identical residue matches

“.” indicates similar residue matches