Chapter 17:
From Gene to Protein
4. Translation
3. The Genetic Code
2. Transcription
1. Overview of Gene Expression
5. Mutations
1. Overview of Gene Expression
Chapter Reading – pp. 334-337
How are Genes related to DNA?
Genes are segments of DNA that code for a
particular protein (or RNA molecule)
• the human genome contains ~3 billion base
pairs (bps) and ~25,000 genes
• most genes encode proteins
• when we talk about “genes” we will focus on those
that express proteins
• the gene products of some genes are RNA
molecules that play a variety of roles in cells
Minimal medium
No growth:
Mutant cells
cannot grow
and divide
Growth:
Wild-type
cells growing
and dividing
EXPERIMENT RESULTS
CONCLUSION
Classes of Neurospora crassa
Wild type Class I mutants Class II mutants Class III mutants
Minimalmedium(MM)(control)
MM ornithine
MM citrulline
Co
nd
itio
n
MM arginine(control)
Summaryof results
Can grow withor without anysupplements
Can grow onornithine,citrulline, orarginine
Can grow onlyon citrulline orarginine
Require arginineto grow
Wild type
Class I mutants
(mutation in
gene A)
Class II mutants
(mutation in
gene B)
Class III mutants
(mutation in
gene C)
Gene
(codes for
enzyme)
Gene A
Gene B
Gene C
Precursor Precursor Precursor Precursor
Enzyme A Enzyme A Enzyme A Enzyme A
Enzyme B Enzyme B Enzyme B Enzyme B
Enzyme C Enzyme C Enzyme C Enzyme C
Ornithine Ornithine Ornithine Ornithine
Citrulline Citrulline Citrulline Citrulline
Arginine Arginine Arginine Arginine
• these classic
experiments led to
the “one gene-
one enzyme”
hypothesis
• we now know that
genes code for
more than just
enzymes
Beadle & Tatum, 1941
Srb & Horowitz, 1944
Gene Expression
The expression of most genes involves two
distinct processes:
1) Transcription of a gene into RNA
• this is essentially creating a “photocopy” of the gene
• occurs in the nucleus
2) Translation of the RNA transcript into protein
• accomplished by ribosomes, in the cytoplasm
• RNA is a nucleic acid very similar to DNA
(RNA uses “U” instead of “T”)
DNA
mRNA
Ribosome
Polypeptide
TRANSCRIPTION
TRANSLATION
TRANSCRIPTION
TRANSLATION
Polypeptide
Ribosome
DNA
mRNA
Pre-mRNARNA PROCESSING
(a) Bacterial cell (b) Eukaryotic cell
Nuclear
envelope
DNA RNA Protein
Prokaryotes vs Eukaryotes
DNAtemplatestrand
TRANSCRIPTION
mRNA
TRANSLATION
Protein
Amino acid
Codon
Trp Phe Gly
5
5
Ser
U U U U U
3
3
53
G
G
G G C C
T
C
A
A
AAAAA
T T T T
T
G
G G G
C C C G G
DNA
molecule
Gene 1
Gene 2
Gene 3
C C
From DNA to RNA to Protein
• gene expression ends at transcription for genes encoding
RNA gene products
Comparison of DNA & RNA
• sugar = Ribose
• single-stranded
• A, C, G & U (uracil)
RNA DNA
• sugar = Deoxyribose
• double-stranded
• A, C, G & T (thymine)
2. Transcription
Chapter Reading – pp. 340-345
Stages of
Transcription
INITIATIONRNA polymerase binds
to the promoter of a
gene, separates DNA
ELONGATIONRNA polymerase makes
RNA complementary to
the template strand
TERMINATIONRNA polymerase stops
transcribing when end of
gene is reached
Promoter
RNA polymerase
Start pointDNA
53
Transcription unit
35
Elongation
53
35
Nontemplate strand of DNA
Template strand of DNARNAtranscript
UnwoundDNA
2
353
53
RewoundDNA
RNAtranscript
5
Termination3
3
5
5Completed RNA transcript
Direction of transcription (“downstream”)
53
3
Initiation1
Transcription initiation
complex forms
3
DNA
PromoterNontemplate strand
53
53
53
Transcription
factors
RNA polymerase II
Transcription factors
53
5
3
53
RNA transcript
Transcription initiation complex
53
TATA box
T
T T T T T
A A A AA
A A
T
Several transcription
factors bind to DNA
2
A eukaryotic promoter1
Start point Template strand
Initiation of
Eukaryotic
Transcription
Eukaryotic promoters
contain a short
sequence called a
TATA box.
With the help of an
array of transcription
factors, RNA
polymerase binds to
the promoter and
starts transcription.
Nontemplate
strand of DNA
RNA nucleotides
RNA
polymerase
Template
strand of DNA
3
35
5
5
3
Newly made
RNA
Direction of transcription
A A A
A
T
TTT G
C
C C
G
C CC A AU
end
ELONGATION
In self-termination, the transcription of DNA
terminator sequences cause the RNA to fold,
loosening the grip of RNA polymerase on the DNA.
In enzyme-dependent termination, a termination
enzyme pushes between RNA polymerase and
the DNA, releasing the polymerase.
Termination of transcription
RNA transcript released
RNA polymerase
released
Rho termination
protein
3
5
Rho protein moves
along RNA
3
C-G rich
stem-loop
Termination of Transcription
• triggered by stem/loop structure in RNA or termination
factors such as the Rho protein (prokaryotes)
Protein-coding
segmentPolyadenylation
signal5 3
35 5Cap UTRStart
codon
G P P P
Stop
codon UTR
AAUAAA
Poly-A tail
AAA AAA…
Primary RNA TranscriptsNewly made RNA molecules in eukaryotes are called
primary transcripts because they need to be
processed before they can carry out their function:
mRNA 1o transcript
• a 5’ cap
Messenger (mRNA) primary transcripts require the
following modifications:
• a poly-A tail
• splicing out of introns
5 Exon Intron Exon
5 CapPre-mRNACodonnumbers
130 31104
mRNA 5 Cap
5
Intron Exon
3 UTR
Introns cut out and
exons spliced together
3
105
146
Poly-A tail
Coding
segment
Poly-A tail
UTR1146
RNA Splicing
The coding region of the primary RNA transcript
contains intervening sequences called introns that
need to be removed or “spliced out”.
The regions that are retained are called exons which
after splicing form a continuous coding region.
RNA transcript (pre-mRNA)5
Exon 1
Protein
snRNA
snRNPs
Intron Exon 2
Other proteins
Spliceosome
5
Spliceosome
componentsCut-out
intronmRNA5
Exon 1 Exon 2
How is
Splicing
Carried
Out?
Spliceosomes are
structures made of
protein and snRNA
snRNA in
spliceosome base
pairs with ends of
intron sequences
resulting in their
being “spliced out”
Exons and Protein Structure
Exons tend to
code for distinct
substructures
called domains
in proteins.
Exon shuffling is
thought to be an
evolutionary
process by
which new
genes are made.
GeneDNA
Exon 1 Exon 2 Exon 3Intron Intron
Transcription
RNA processing
Translation
Domain 3
Domain 2
Domain 1
Polypeptide
Various Roles of RNA Transcripts
1) messenger RNA (mRNA)
• RNA copy of a gene that encodes a polypeptide
2) ribosomal RNA (rRNA)
• RNA that is a structural component of ribosomes
3) transfer RNA (tRNA)
• delivery of “correct”
amino acids to
ribosomes during
translation
For some genes, the end-product
is the RNA itself (rRNA, tRNA)
3. The Genetic Code
Chapter Reading – pp. 337-340
How do Genes code for Proteins?
Recall that proteins are linear polymers
made of 20 different amino acids.
Genes need simply to encode the identity of
each amino acid in a given protein!
• i.e., genes must be capable of encoding
20 different amino acids and their order in a
protein
• although DNA contains only 4 different
nucleotides, this is more than sufficient to
specify 20 different amino acids…
Basis of the Genetic Code
Each amino acid in a protein is specified by
3 nucleotide sequences called codons
• there are 64 possible “codon” triplets (4 x 4 x 4)
• more than enough to encode 20 amino acids and the
signal to “stop” or end the protein (TGA, TAA or TAG)
• each of the 20 amino acids is coded for by a
unique set of codons:
e.g. ATG = methionine (start codon)
GGN = glycine
CAA or CAG = glutamine
If the DNA sequence is:5’-CATGCCTGGGCAATAG-3’
3’-GTACGGACCCGTTATC-5’
The mRNA transcript is: 5’-CAUGCCUGGGCAAUAG-3’
The polypeptide is:
*Met-Pro-Gly-Gln (stop)
all proteins begin w/Met
(transcription)
(translation)
The Genetic CodeSecond mRNA base
Fir
st
mR
NA
base (
5en
d o
f co
do
n)
Th
ird
mR
NA
base (
3en
d o
f co
do
n)
UUU
UUC
UUA
CUU
CUC
CUA
CUG
Phe
Leu
Leu
Ile
UCU
UCC
UCA
UCG
Ser
CCU
CCC
CCA
CCG
UAU
UACTyr
Pro
Thr
UAA Stop
UAG Stop
UGA Stop
UGU
UGCCys
UGG Trp
GC
U
U
C
A
U
U
C
C
CA
U
A
A
A
G
G
His
Gln
Asn
Lys
Asp
CAU CGU
CAC
CAA
CAG
CGC
CGA
CGG
G
AUU
AUC
AUA
ACU
ACC
ACA
AAU
AAC
AAA
AGU
AGC
AGA
Arg
Ser
Arg
Gly
ACGAUG AAG AGG
GUU
GUC
GUA
GUG
GCU
GCC
GCA
GCG
GAU
GAC
GAA
GAG
Val Ala
GGU
GGC
GGA
GGGGlu
Gly
G
U
C
A
Met or
start
UUG
G
(a) Tobacco plant expressing a firefly gene gene
(b) Pig expressing a jellyfish
The Genetic Code is UniversalGenes from one species can be expressed in another!
4. Translation
Chapter Reading – pp. 83, 345-354
Polypeptide
Ribosome
tRNA with
amino acid
attached
Amino
acids
tRNA
Anticodon
Codons
U U U UG G G G C
C
G
C
G
5 3
mRNA
Overview of
Translation
Ribosomes facilitate
the production of
polypeptides by:
1) matching codons
in mRNA with
complementary
anticodons in tRNA
2) catalyzing peptide
bonds between
amino acids carried
by tRNAs
tRNA
molecules
Growing
polypeptide Exit tunnel
E PA
Large
subunit
Small
subunit
mRNA5
3
(a) Computer model of functioning ribosome
Exit tunnel Amino end
A site (Aminoacyl-
tRNA binding site)
Small
subunit
Large
subunit
E P AmRNA
E
P site (Peptidyl-tRNA
binding site)
mRNA
binding site
(b) Schematic model showing binding sites
E site
(Exit site)
(c) Schematic model with mRNA and tRNA
5 Codons
3
tRNA
Growing polypeptide
Next amino
acid to be
added to
polypeptide
chain
Ribosome
Structure
Amino acid
attachment
site
3
5
Hydrogen
bonds
Anticodon
(a) Two-dimensional structure (b) Three-dimensional structure
(c) Symbol used in this book
Anticodon Anticodon
3 5
Hydrogen
bonds
Amino acid
attachment
site5
3
A A G
Transfer RNA (tRNA)tRNA anticodon
will base pair with
a complementary
codon in mRNA
Aminoacyl-tRNA
synthetase (enzyme)
Amino acid
P P P Adenosine
ATP
P
P
P
PPi
i
i
Adenosine
tRNA
AdenosineP
tRNA
AMP
Computer model
Amino
acid
Aminoacyl-tRNA
synthetase
Aminoacyl tRNA
(“charged tRNA”)
Aminoacyl-tRNA
Synthetases
Each tRNA
is loaded
with the
correct
amino acid
due to the
action of
these
enzymes.
Initiator
tRNA
mRNA
5
53
Start codonSmall
ribosomal
subunitmRNA binding site
3
Translation initiation complex
5 3
3 U
U
AA G
C
P
P site
i
GTP GDP
Large
ribosomal
subunit
E A
5
Initiation of Translation
• small ribosomal subunit aligns with start codon of mRNA
• initiator tRNAmet and large subunit then join the complex
Amino end ofpolypeptide
mRNA
5
E
Asite
3
E
GTP
GDP P i
P A
E
P A
GTP
GDP P i
P A
E
Ribosome ready fornext aminoacyl tRNA
Psite
Elongation
Cycle of
Translation
Releasefactor
Stop codon
(UAG, UAA, or UGA)
3
5
3
5
Freepolypeptide
2 GTP
5
3
2 GDP 2i
P
Termination of Translation
When a stop codon is reached there is no tRNA
with a complementary anticodon.
Instead a release factor fits in the A site and
catalyzes the dissociation of all components.
Multiple
Ribosomes
translate the
same mRNA
Referred to as
polyribosomes.
Completed
polypeptide
Incoming
ribosomal
subunits
Start of
mRNA
(5 end)
End of
mRNA
(3 end)(a)
Ribosomes
mRNA
(b)0.1 m
Growing
polypeptides
The cap comesoff, and the properlyfolded protein isreleased.
The cap comesoff, and the properlyfolded protein isreleased.
Hollowcylinder
Cap
Chaperonin(fully assembled)
Polypeptide
Steps of ChaperoninAction:
An unfolded poly-peptide enters thecylinder from one end.
1
2 3The cap attaches, causing thecylinder to change shape insuch a way that it creates ahydrophilic environment forthe folding of the polypeptide.
The cap comesoff, and the properlyfolded protein isreleased.
Correctlyfoldedprotein
Ensuring Proper Protein Structure (pg. 83)
Chaperonins are large protein structures that assist
newly made polypeptides to ensure they fold properly.
• capture improperly folded proteins inside its cavity, forcing
them to unfold and hopefully refold correctly
Ribosome
mRNA
Signalpeptide
SRP
1
SRPreceptorprotein
Translocationcomplex
ERLUMEN
2
3
45
6
Signalpeptideremoved
CYTOSOL
Protein
ERmembrane
Translation in the ER
Proteins destined for the “secretory pathway”
have a signal peptide at the N-terminus that
targets the protein to the ER lumen.
Gene Expression in Prokaryotes
Transcription and
translation are
not segregated in
prokaryotes.
Since prokaryotic
RNA transcripts
don’t need to be
processed,
translation can
begin before
transcription is
finished!
RNA polymerase
DNAmRNA
Polyribosome
RNA
polymeraseDNA
Polyribosome
Polypeptide
(amino end)
mRNA (5 end)
Ribosome
0.25 mDirection of
transcription
TRANSCRIPTIONDNA
RNApolymerase
Exon
RNAtranscript
RNAPROCESSING
NUCLEUS
Intron
RNA transcript(pre-mRNA)
Aminoacyl-tRNA synthetase
AMINO ACIDACTIVATION
Aminoacid
tRNA
3
GrowingpolypeptidemRNA
Aminoacyl(charged)tRNA
Anticodon
Ribosomalsubunits
A
AE
TRANSLATION
CYTOPLASM
P
E
Codon
Ribosome
5
3
Summary
of Gene
Expression
5. Mutation
Chapter Reading – pp. 355-357
Mutations
A mutation is any change in DNA sequence:
• change of one nucleotide to another
• insertion or deletion of nucleotides or DNA fragments
• inversion or recombination of DNA fragments
What causes mutations?
• errors in DNA replication or DNA repair
• chemical mutagenesis
• high energy electromagnetic radiation
• UV light, X-rays, gamma rays
Sickle-cell Anemia
• a single nucleotide change causes a single amino
acid change resulting in a malformed protein
Wild-type hemoglobin
Wild-type hemoglobin DNA
3
3
35
5 3
35
5
553
mRNA
A AG
C T T
A AG
mRNA
Normal hemoglobin
Glu
Sickle-cell hemoglobin
Val
A
A
AUG
G
T
T
Sickle-cell hemoglobin
Mutant hemoglobin DNA
C
Types of Mutations
Silent mutations:
• have no effect on amino acid specification
Missense mutations:
• result in the change of a single amino acid
Nonsense mutations:
• convert a codon specifying an amino acid to a stop codon
• results in premature truncation of a protein
Insertion/deletion mutations:
• cause a shift in the reading frame of the gene
• all codons downstream of insertion/deletion will be incorrect
Silent MutationsWild type
DNA template strand
mRNA5
5
Protein
Amino end
Stop
Carboxyl end
3
3
3
5
Met Lys Phe Gly
A instead of G
(a) Nucleotide-pair substitution: silent
StopMet Lys Phe Gly
U instead of C
A
A
A A
A A A A
A AT
T T T T T
T T TT
C C C C
C
C
G G G G
G
G
A
A A A AG GGU U U U U
5
3
3
5A
A A
A A A A
A AT
T T T T T
T T TT
C C C C
G G G G
A
A
A G A A A AG GGU U U U U
T
U 35
Missense MutationsWild type
DNA template strand
mRNA5
5
Protein
Amino end
Stop
Carboxyl end
3
3
3
5
Met Lys Phe Gly
T instead of C
(a) Nucleotide-pair substitution: missense
StopMet Lys Phe Ser
A instead of G
A
A
A A
A A A A
A AT
T T T T T
T T TT
C C C C
C
C
G G G G
G
G
A
A A A AG GGU U U U U
5
3
3
5A
A A
A A A A
A AT
T T T T T
T T TT
C C T C
G
G
GA
A G A A A AA GGU U U U U 35
A C
C
G
Nonsense MutationsWild type
DNA template strand
mRNA5
5
Protein
Amino end
Stop
Carboxyl end
3
3
3
5
Met Lys Phe Gly
A instead of T
(a) Nucleotide-pair substitution: nonsense
Met
A
A
A A
A A A A
A AT
T T T T T
T T TT
C C C C
C
C
G G G G
G
G
A
A A A AG GGU U U U U
5
3
3
5A
A
A A A A
A AT
T A T T T
T T TT
C C C
G
G
GA
A G U A A AGGU U U U U 35
C
C
G
T instead of C
C
GT
U instead of A
G
Stop
Wild type
DNA template strand
mRNA5
5
Protein
Amino end
Stop
Carboxyl end
3
3
3
5
Met Lys Phe Gly
A
A
A A
A A A A
A AT
T T T T T
T T TT
C C C C
C
C
G G G G
G
G
A
A A A AG GGU U U U U
(b) Nucleotide-pair insertion or deletion: frameshift causing
immediate nonsense
Extra A
Extra U
5
3
5
3
3
5
Met
1 nucleotide-pair insertion
Stop
A C A A GT T A TC T A C G
T A T AT G T CT GG A T GA
A G U A U AU GAU G U U C
A T
A
AG
Frameshift – Insertion/Deletion
Can cause a premature
stop codon…
DNA template strand
mRNA5
5
Protein
Amino endStop
Carboxyl end
3
3
3
5
Met Lys Phe Gly
A
A
A A
A A A A
A AT
T T T T T
T T TT
C C C C
C
C
G G G G
G
G
A
A A A AG GGU U U U U
(b) Nucleotide-pair insertion or deletion: frameshift causing
extensive missense
Wild type
missing
missing
A
U
A A AT T TC C A T TC C G
A AT T TG GA A ATCG G
A G A A GU U U C A AG G U 3
5
3
3
5
Met Lys Leu Ala
1 nucleotide-pair deletion
5
Frameshift – Insertion/Deletion
…or
extended
shift in
reading
frame…
DNA template strand
mRNA5
5
Protein
Amino end
Stop
Carboxyl end
3
3
3
5
Met Lys Phe Gly
A
A
A A
A A A A
A AT
T T T T T
T T TT
C C C C
C
C
G G G G
G
G
A
A A A AG GGU U U U U
(b) Nucleotide-pair insertion or deletion: no frameshift, but one
amino acid missing
Wild type
AT C A A A A T TC C G
T T C missing
missing
Stop
5
3
3
5
35
Met Phe Gly
3 nucleotide-pair deletion
A GU C A AG GU U U U
T GA A AT T TT CG G
A A G
Frameshift – Insertion/Deletion
…or the
addition/
deletion of
a single
amino acid.
Must be a
multiple
of 3!
Key Terms for Chapter 17
• transcription, promoter, TATA box, RNA polymerase
• primary transcript, mRNA, tRNA, rRNA, snRNA
• 5’ cap, poly-A tail, intron, exon, splicing, spliceosome
• rho protein, stem-loop, codon, anti-codon, translation
• mutation: substitution, deletion, insertion
• silent, missense, nonsense mutations
• aminoacyl tRNA synthetase, polyribosome, signal
peptide, release factor, chaperonin
• reading frame, frameshift
Relevant
Chapter
Questions 1-9, 11
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