D1.

2 Protein synthesis
Continuity and change—Molecules
Standard level and higher level: 3
hours
Additional higher level: 3 hours
D1.2 Protein synthesis
Continuity and change—Molecules
Standard level and higher level: 3 hours
Additional higher level: 3 hours

SL and HL
D1.2.1—Transcription as the synthesis of RNA using a DNA template
Students should understand the roles of RNA polymerase in this process.
D1.2.2—Role of hydrogen bonding and complementary base pairing in transcription
Include the pairing of adenine (A) on the DNA template strand with uracil (U) on the RNA strand.
D1.2.3—Stability of DNA templates
Single DNA strands can be used as a template for transcribing a base sequence, without the DNA base
sequence changing. In somatic cells that do not divide, such sequences must be conserved throughout
the life of a cell.
D1.2.4—Transcription as a process required for the expression of genes
Limit to understanding that not all genes in a cell are expressed at any given time and that transcription,
being the first stage of gene expression, is a key stage at which expression of a gene can be switched on
and off.
D1.2.5—Translation as the synthesis of polypeptides from mRNA
The base sequence of mRNA is translated into the amino acid sequence of a polypeptide.
D1.2.6—Roles of mRNA, ribosomes and tRNA in translation
Students should know that mRNA binds to the small subunit of the ribosome and that two tRNAs can bind
simultaneously to the large subunit.
D1.2.7—Complementary base pairing between tRNA and mRNA
Include the terms “codon” and “anticodon”.
D1.2.8—Features of the genetic code
Students should understand the reasons for a triplet code. Students should use and understand the terms
“degeneracy” and “universality”.
D1.2 Protein synthesis
Continuity and change—Molecules
Standard level and higher level: 3 hours
Additional higher level: 3 hours

D1.2.9—Using the genetic code expressed as a table of mRNA codons

Students should be able to deduce the sequence of amino acids coded by an mRNA strand.
D1.2.10—Stepwise movement of the ribosome along mRNA and linkage of amino acids by peptide
bonding to the growing polypeptide chain
Focus on elongation of the polypeptide, rather than on initiation and termination.
D1.2.11—Mutations that change protein structure
Include an example of a point mutation affecting protein structure.
D1.2 Protein synthesis
Continuity and change—Molecules
Standard level and higher level: 3 hours
Additional higher level: 3 hours
Additional higher level
D1.2.12—Directionality of transcription and translation
Students should understand what is meant by 5' to 3' transcription and 5' to 3' translation.
D1.2.13—Initiation of transcription at the promoter
Consider transcription factors that bind to the promoter as an example. However, students are not required to
name the transcription factors.
D1.2.14—Non-coding sequences in DNA do not code for polypeptides
Limit examples to regulators of gene expression, introns, telomeres and genes for rRNAs and tRNAs in
eukaryotes.
D1.2.15—Post-transcriptional modification in eukaryotic cells
Include removal of introns and splicing together of exons to form mature mRNA and also the addition of 5'
caps and 3' polyA tails to stabilize mRNA transcripts.
D1.2.16—Alternative splicing of exons to produce variants of a protein from a single gene
Students are only expected to understand that splicing together different combinations of exons allows
one gene to code for different polypeptides. Specific examples are not required.
D1.2.17—Initiation of translation
Include attachment of the small ribosome subunit to the 5' terminal of mRNA, movement to the start
codon, the initiator tRNA and another tRNA, and attachment of the large subunit. Students should
understand the roles of the three binding sites for tRNA on the ribosome (A, P and E) during elongation.
D1.2.18—Modification of polypeptides into their functional state
Students should appreciate that many polypeptides must be modified before they can function. The
examples chosen should include the two-stage modification of pre-proinsulin to insulin.
D1.2.19—Recycling of amino acids by proteasomes
Limit to the understanding that sustaining a functional proteome requires constant protein breakdown
and synthesis.
 Genes to Proteins

The central dogma of molecular biology has stated that genetic

information, encoded in DNA, is transcribed into molecules of
RNA, which are then translated into the amino acid sequences
that make up proteins. The nature of a protein determines its
role in the cell. Amin
o acid

Structural?
tRNA
Regulatory?
Reverse
transcription
Contractile?

Immunological?
Transcription Translation
Transport?
Protein
DNA mRNA Catalytic?
 Prokaryotic Gene Expression
The process of transferring the information encoded in a gene to
its functional gene product is called gene expression.
– The simple “one-gene protein” model is supported by studies of prokaryotic genomes,
where much of the DNA comprises protein-coding genes.

Gene for a protein

DNA
Transcription

Gene regulation
RNA transcript Translation

The DNA of prokaryotes

consists almost entirely
of protein-coding genes Protein
and their regulatory
sequences Structural and regulatory functions
 Eukaryotic Gene Expression
DNA gene containing both intronic and exonic DNA
Gene regulation

Gene regulation
Primary RNA Transcription
transcript

Splicing
Intronic RNA
Assembled
exonic RNA
processing

processing

mRNA
Translation microRNAs

Non-coding
RNA
Roles in genomic Role in the timing of
Diverse development
functions regulation
 Transcription
DNA

Free nucleotides
Single-armed
used to construct
chromosome
the mRNA strand
as found in
non-dividing
cell
A mRNA strand is formed
RNA polymerase
using the DNA molecule enzyme

as the template. Template strand of

DNA contains the
information for the
Free nucleotides with

D
construction of a

ire of esi
ct
bases complementary to

sy
functional mRNA

io
nt

n
h
product (e.g. a
the DNA are joined

s
Coding protein)

together by the enzyme strand

RNA polymerase. The two strands
of DNA coil up Formation of a single strand of
into a double mRNA that is complementary to
helix
the template strand (therefore
the same “message” as the
coding strand)
 Transcription

Initiation. RNA polymerase recognizes the promoter and binds to

the DNA. A promoter is a section of the DNA that initiates gene
transcription. A promoter is typically between 100 and 1,000
bases long and is located adjacent to genes on chromosomes.
 Transcription

The base sequence of a promoter allows RNA polymerase to bind

along with a variety of proteins that act as transcription factors.
 Transcription
After RNA polymerase binds to the promoter (short sequence of
bases that is not transcribed), the DNA strands unwind, and the
polymerase initiates RNA synthesis at the start point on the
template strand.
 Transcription

Activators .- sequences within the promoter (enhancer) allow

transcription factors to bind that encourage binding of RNA
polymerase, leading to transcription
 Transcription

Repressors.- sequences within the promoter (silencers) allow

transcription factors to bind that prevent binding of RNA
polymerase, so transcription of a gene is prevented.
 Transcription

Elongation. The RNA polymerase moves downstream,

unwinding the DNA and elongating the RNA transcript 5¢ ® 3 ¢.
In the wake of transcription, the DNA strands re-form a double
helix.
 Transcription

Termination. Eventually, the RNA transcript is released, and

the RNA polymerase detaches from the DNA.
Synthesis of mRNA
 Sense and Antisense strands

The DNA strand that carries the genetic code is called the
sense strand (or the coding strand). The other strand is
called the antisense strand (or the template strand). The
sense strand has the same sequence as the newly
transcribed mRNA, except with thymine in place of uracil
 Alteration of mRNA Ends

In eukaryotes, the locations for transcription and

translation are separated by the nuclear membrane.
This allows for post-transcriptional modification of
the mRNA. The first product of transcription is pre-
mRNA
 Alteration of mRNA Ends

Each end of a pre-mRNA molecule is modified

in a particular way
– The 5¢ end receives a modified nucleotide cap
– The 3¢ end gets a poly-A tail

A modified guanine nucleotide 50 to 250 adenine nucleotides

added to the 5¢ end added to the 3¢ end
TRANSCRIPTION DNA

RNA PROCESSING Pre-mRNA Protein-coding segment Polyadenylation signal

5¢ 3¢
mRNA
G P P P AAUAAA AAA…AAA
Ribosome
TRANSLATION
Start codon Stop codon
5¢ Cap 5¢ UTR 3¢ UTR Poly-A tail
Polypeptide
 Alteration of mRNA Ends

The 5ʹ cap and the poly (A) tail both stabilize the ends of the
mRNA by protecting them from digestion by nuclease enzymes.
They also encourage further modification of the original
transcript
A modified guanine nucleotide 50 to 250 adenine nucleotides
added to the 5¢ end added to the 3¢ end
TRANSCRIPTION DNA

RNA PROCESSING Pre-mRNA Protein-coding segment Polyadenylation signal

5¢ 3¢
mRNA
G P P P AAUAAA AAA…AAA
Ribosome
TRANSLATION
Start codon Stop codon
5¢ Cap 5¢ UTR 3¢ UTR Poly-A tail
Polypeptide
 Split Genes and RNA Splicing
Eukaryotic genes that code for polypeptides contain two types
of sequences
Exons—coding sequences that are expressed by translation into
the amino acid sequence.
Introns—intervening sequences that are not expressed.Introns
are mostly between 20 and 200 nucleotides long. The mean
number of introns per gene in humans is 7.8 and the mean
number of exons is 8.8.
5¢ Exon Intron Exon Intron Exon 3¢
TRANSCRIPTION DNA Pre-mRNA 5¢ Cap Poly-A tail
1 30 31 104 105 146
RNA PROCESSING Pre-mRNA

mRNA Coding Introns cut out and

segment exons spliced together
Ribosome
TRANSLATION

Polypeptide mRNA 5¢ Cap Poly-A tail

1 146
3¢ UTR 3¢ UTR
 Split Genes and RNA Splicing

RNA splicing
– Removes introns (non-coding) and joins exons
(coding)

5¢ Exon Intron Exon Intron Exon 3¢

TRANSCRIPTION DNA Pre-mRNA 5¢ Cap Poly-A tail
1 30 31 104 105 146
RNA PROCESSING Pre-mRNA

mRNA Coding Introns cut out and

segment exons spliced together
Ribosome
TRANSLATION

Polypeptide mRNA 5¢ Cap Poly-A tail

1 146
3¢ UTR 3¢ UTR
 Split Genes and RNA Splicing

RNA splicing is carried out by spliceosomes

RNA transcript (pre-mRNA)
5¢
Exon 1 Intron Exon 2
Protein
1 Other proteins
snRNA
snRNPs
Spliceosome

2 5¢

Spliceosome
components
Cut-out
mRNA intron
3 5¢
Exon 1 Exon 2
 Split Genes and RNA Splicing

Alternative splicing can occur with genes that produce multiple

proteins, which means that some exons may also be removed
during splicing, thus producing different polypeptides
 Split Genes and RNA Splicing

In mammals, the protein tropomyosin is encoded by a gene

that has 11 exons. Tropomyosin pre-mRNA is spliced differently
in different tissues resulting in fve different forms of the
protein. For example, in skeletal muscle, exon 2 is missing from
the mRNA; in smooth muscle, exons 3 and 10 are missing from
the mRNA.
Non-coding sequences in DNA do
not code for polypeptides
There are also base sequences in genomes that do not code for
polypeptides, but have other functions. Five examples of base
sequences that have a function other than coding for a
polypeptide are

• base sequences that can be transcribed to produce tRNA

• base sequences that can be transcribed to produce ribosomal RNA (rRNA)
• base sequences used in the regulation of gene expression such as
promoters (the binding sites for RNA polymerase), enhancers and silencers
• telomeres—the structures that form the ends of chromosomes in
eukaryotes
• introns—base sequences that are edited out of mRNA during post-
transcriptional modification
 Movement of mRNA
In eukaryotic cells, the two
main steps in protein
synthesis occur in separate
compartments: transcription
in the nucleus and Ribosomes
translation in the cytoplasm. mRNA

– mRNA moves out of

the nucleus, to the
cytoplasm, through pores in
the nuclear membrane.

In prokaryotic cells, there is

no nucleus, and the
chromosome is in direct Nuclear pore through
contact with the cytoplasm, which the mRNA

and protein synthesis can passes

into the cytoplasm
begin even while the DNA is
being transcribed.
Nucleus Cytoplasm
 Overview of transcription and translation

Translation is the process of building a polypeptide chain

from amino acids, guided by the sequence of codons
(sequence of three bases) on the mRNA.
 mRNA Codes for Amino Acids
Read second
letter here Second Letter Read third
letter here
Read first
letter here U C A G
UUU Phe UCU Ser UAU Tyr UGU Cys U
UUC Phe UCC Ser UAC Tyr UGC Cys C
U UUA Leu UCA Ser UAA STOP UGA STOP A
UUG Leu UCG Ser UAG STOP UGG Try G
CUU Leu CCU Pro CAU His CGU Arg U
CUC Leu CCC Pro CAC His CGC Arg C
C
First Letter

Third Letter
CUA Leu CCA Pro CAA Gln CGA Arg A
CUG Leu CCG Pro CAG Gln CGG Arg G
AUU Iso ACU Thr AAU Asn AGU Ser U
AUC Iso ACC Thr AAC Asn AGC Ser C
A AUA Iso ACA Thr AAA Lys AGA Arg A
AUG Met ACG Thr AAG Lys AGG Arg G
GUU Val GCU Ala GAU Asp GGU Gly U
GUC Val GCC Ala GAC Asp GGC Gly C
G GUA Val GCA Ala GAA Glu GGA Gly A
GUG Val GCG Ala GAG Glu GGG Gly G
Features of the
genetic code
Two notable features of the genetic code are degeneracy and
universality. "Degenerate" refers to redundancy in the genetic
code. Amino acids, the building blocks of proteins, are encoded
by codons of three nucleotide bases. Some amino acids are
encoded by more than one codon.

A mutation altering one base in a codon is unlikely to alter the

amino acid structure of the encoded protein, because the
codon is likely to still encode the same amino acid. This makes
the genetic code more fault-tolerant to point mutations.
 Translation

Structures involved in
translation:
– Messenger RNA molecules (mRNA) The speckled appearance of the rough
carries endoplasmic reticulum is the result of
the code from the DNA that will be ribosomes bound to the membrane
translated surface.

into an amino acid sequence.

– Transfer RNA molecules (tRNA) mRNA

transport amino acids to their correct

position on the mRNA strand.
– Ribosomes provide the environment for
tRNA attachment and amino acid Ribosomes
linkage. tRNA

– Amino acids from which the

polypeptides Amino acids
are constructed.
 Ribosomes & tRNA
Ribosome Ribosome

– Comprises two subunits in which

there are grooves where the mRNA
strand and polypeptide chain fit in.
– The ribosomal subunits are Large Small
constructed of protein and subunit subunit
ribosomal RNA (rRNA).
Amino acid attachment
– The subunits form a functional unit site
only when they attach to a mRNA Ribosome
attachment
molecule. point

tRNA molecule
– There is a specific tRNA molecule
and anticodon for each type of
codon.
The 3-base sequence of
– The anticodon is the site of the 3- the anticodon is
base sequence that 'recognizes' and complementary to the
matches up with the codon on the Anticodon
codon on the mRNA
mRNA molecule. molecule
Transfer RNA molecule
 The Structure and Function of Transfer RNA

A
C
C
A tRNA molecule consists of a single RNA strand that
is only about 80 nucleotides long
3¢
Amino acid A
C
attachment site C
A 5¢
The four base-paired regions and C
G
G
C
three loops are characteristic of C
U
U
G
G
A
all tRNAs, as is the base sequence U C
A
A
U
U
* C A C A G UA A G *
of the amino acid attachment site G
C G U G U *
* C U C
C G A G G
*
* U C
at the 3¢ end. The anticodon * A G G
*
* G AG C Hydrogen
triplet is unique to each tRNA G
U
C
A bonds
* G
type that pairs with a specic A * A
C
* U
codon of mRNA. A
A G

Anticodon
 Activation of Transfer RNA

Amino acid Aminoacyl-tRNA

synthetase (enzyme)

1 Active site binds the

A specific enzyme
P P P Adenosine
amino acid and ATP.
ATP

called an 2 ATP loses two P groups

and joins amino acid as AMP.
aminoacyl-tRNA P Adenosine

synthetase
Pyrophosphate P Pi

Pi

– Joins each
Pi
Phosphates
tRNA
amino acid 3 Appropriate
tRNA covalently
to the Bonds to amino
Acid, displacing P Adenosine

correct tRNA AMP.

AMP

4 Activated amino acid

is released by the enzyme.

Aminoacyl tRNA
(an “activated
amino acid”)
Stages of Translation
 Translation: Initiation

The first stage of translation involves the assembly of the three

components that carry out the process (mRNA, tRNA, ribosome)

Initiator
tRNA Activated
Thr-tRNA

Small ribosomal
unit attaches

Large ribosomal unit

attaches to form a functional
ribosomal protein-
synthesizing complex

mRNA

P A
site site Ribosomes move in this direction
Ribosome
 Translation: Initiation
Initiation
• 5’ end of mRNA binds to the small subunit of the ribosome
• initial mRNA codon = AUG = start codon
• tRNA with anticodon: UAC binds to mRNA AUG codon by complementary
base pairing, methionine attached to tRNA 3’ terminal
• large ribosomal subunit binds, completing ribosomal structure, and
producing two ribosomal binds sites: P site & A site
Initiator
tRNA Activated
Thr-tRNA

Small ribosomal
unit attaches

Large ribosomal unit

attaches to form a functional
ribosomal protein-
synthesizing complex

mRNA

P A
site site Ribosomes move in this direction
Ribosome
The ribosome has P site (Peptidyl-tRNA
binding site)
three binding A site (Aminoacyl-
tRNA binding site)

sites for tRNA E site

(Exit site)

The P site , The Large

subunit
E P A
A site, The E site
mRNA
binding site Small
subunit

(b) Schematic model showing binding sites. A ribosome has an mRNA binding site
and three tRNA binding sites, known as the A, P, and E sites. This schematic
ribosome will appear in later diagrams.
 Elongation and Translocation

In the Elongation stage of translation, amino

acids are added one by one by tRNAs as the
ribosome moves along the mRNA (5¢ ® 3 ¢).

Activated
Tyr-tRNA

Growing
polypeptide

Unloaded
Thr-tRNA

mRNA
5’
P A
site site
There are three steps:
• The correct tRNA binds to the A site on the
ribosome.
• A peptide bond forms between adjacent amino
acids.
• Translocation .- The tRNA at the P site is
released. The tRNA at the A site, now attached
to the growing polypeptide, moves to the P site
and the ribosome advances by one codon Activated
Tyr-tRNA

Growing
polypeptide

Unloaded
Thr-tRNA

mRNA
5’
P A
site site
 Translation: Termination

The final stage of protein

synthesis Termination occurs
when the ribosome reaches
a stop codon.
Completed
polypeptide Completed
polypeptide
is released

Release factor
 Translation: Termination
When ribosomal A-site
reaches a stop codon, no
tRNA has a complementary
anticodon
• when a ribosome
encounters a stop
codon, a release
factor binds to the
stop codon

Release factor protein binds Completed

polypeptide Completed
to ribosomal A-site stop polypeptide
codon and the polypeptide is released
and mRNA are released
large and small ribosomal
Release factor
subunits separate
 Overview of Translation Polypeptide chain in an
Activating advanced stage of
synthesis
Lys-tRNA

Activated
Tyr-tRNA

Growing
polypeptide

Unloaded
Thr-tRNA

Start
codon
Ribosome

mRNA

Ribosomes moving in this direction

 Polysomes

Polysomes are several ribosomes used to translate an

mRNA at the same time, forming what is called a polysome.
More than one ribosome can translate an mRNA at one
time, making it possible to produce many polypeptides
simultaneously from a single mRNA.
• In prokaryotes, several ribosomes can attach themselves to the
growing mRNA chains to form a polysome while the mRNA chains
are still attached to the DNA.

• In eukaryotes, the mRNA detaches from the DNA and is then

transported through pores in the nuclear envelope to the ribosomes
in the cytoplasm. Once in the cytosol, eukaryote mRNA can also
form polysomes.
Polysomes
Modification of polypeptides
after translation
• Removal of the amino acid methionine from the 5ʹ end of
the polypeptide, where it was placed during initiation of
translation.

• Changes to the side chains of amino acids in the

polypeptide. For example, phosphorylation or addition of
carbohydrate molecules, can produce over 100 chemically
different amino acids in proteins.

• Folding the polypeptide and making intramolecular

interactions to stabilize the tertiary structure. For example,
disulfide bonds are formed between cysteines that come close
together due to folding.
Modification of polypeptides
after translation

• Conversion of propeptides into mature peptides by removal

of part of the polypeptide chain.

• Combining two or more polypeptides into the quaternary

structure proteins.

• Combining non-polypeptide components into the

quaternary structure of conjugated proteins.
Modification of
preproinsulin to insulin

The insulin gene (INS) on

chromosome 11 is
transcribed in pancreatic
beta cells to produce
mRNA copies.

The mRNA is translated by

ribosomes on the rough
endoplasmic reticulum.
The polypeptide produced
is preproinsulin and has
110 amino acids
Modification of
preproinsulin to insulin
Preproinsulin passes into
the lumen of the rough
endoplasmic reticulum.
Here a sequence of 24
amino acids (called the
signal peptide) is removed
from the N-terminal by a
protease enzyme. A chain of
86 amino acids remains
which is called proinsulin.
Proinsulin becomes folded
with three disulfide bonds
stabilizing the tertiary
structure.
Modification of
preproinsulin to insulin
Peptide bonds are broken at
two positions in proinsulin by
proteases. This causes a
sequence of 33 amino acids to
be excised and leaves two
separate chains: the A-chain
with 21 amino acids and the B-
chain, initially with 32 amino
acids. These two chains are
held together by the disulfide
bonds. A pair of amino acids is
removed from the C-terminal
of the B-chain by a protease, to
yield mature insulin with a
total of 51 amino acids.
Recycling of amino acids
by proteasomes
Most of the proteins produced by translation have a relatively
short life in the cell. There are many reasons for this.

• The cell’s activities may change, and a protein is no longer

needed. An example of this is when a cell moves on from one
stage in the cell cycle to the next one.

• The structure of many proteins is quite easily changed by

free radicals or other reactive chemicals. If this happens the
protein can no longer carry out its function.

• The protein could become misfolded or denatured in other

ways.
Recycling of amino acids
by proteasomes
Proteins that no longer have a function or are unable to carry
out their function are broken down by proteasomes.
Recycling of amino acids
by proteasomes
Proteins identified for degradation are tagged with a chain of
small proteins called ubiquitin. This acts as a signal for
proteasomes that these proteins should be digested.
Recycling of amino acids
by proteasomes
Ubiquitinated proteins are recognized by the regulatory
subunits at either end of the proteasome. They are then
unfolded and fed into the central chamber by a subunit that
uses ATP as a source of energy.
Recycling of amino acids
by proteasomes
Active sites of multiple proteases face the central chamber
and break proteins down into short chains of amino acids
which pass out of the proteasome.