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From gene to protein
Our plan:
• Overview gene expression
• Walk through the process– Review structure and function of DNA– Transcription – Translation
• Gene expression and mutations
Gene expression- the process by which DNA directs the synthesis of proteins
Overview
DNA mRNA ProteinTranscription Translation
Overview: Cell as a city
DNA: Blueprint for the cityNucleus: City HallNuclear envelope: Fence around City Hall
Overview: Cell as a city
DNA mRNA ProteinTranscription Translation
•The blueprint cannot leave City Hall•Photocopies of the blueprint can be taken out into the city•Architects and builders translate the blue print into the city’s infrastructure
•The DNA remains in the nucleus•Transcription generates mobile RNA transcripts using DNA as a template•The RNA sequence can be translated into a protein
Overview
DNA mRNA ProteinTranscription Translation
DNA
Hydrogen bond 3 end
5 end
3.4 nm
0.34 nm3 end
5 end(b) Partial chemical
structure(a) Key features of DNA structure
1 nm
Genes- discrete units of hereditary information consisting of a specific nucleotide sequence of DNA
Overview
DNA mRNA ProteinTranscription Translation
TranscriptionTranscription-DNA guides the production of RNA
Takes place in three phases:•Initiation•Elongation•Termination
TranscriptionTranscription-DNA guides the production of RNA
Takes place in three phases:•Initiation•Elongation•Termination
•RNA polymerase binds to a promoter in the DNA (contains a signal sequence and at start point)•The DNA is separated and unwound•Transcription begins
-
A eukaryotic promoterincludes a TATA box
3
1
2
3
Promoter
TATA box Start point
Template
TemplateDNA strand
535
Transcriptionfactors
Several transcription factors mustbind to the DNA before RNApolymerase II can do so.
5533
Additional transcription factors bind tothe DNA along with RNA polymerase II,forming the transcription initiation complex.
RNA polymerase IITranscription factors
55 53
3
RNA transcript
Transcription initiation complex
TranscriptionTranscription-DNA guides the production of RNA
Takes place in three phases:•Initiation•Elongation•Termination
Elongation
RNApolymerase
Nontemplatestrand of DNA
RNA nucleotides
3 end
Direction oftranscription(“downstream”) Template
strand of DNANewly madeRNA
3
5
5
•RNA polymerase moves along the template strand•Untwists the helix •Adds complimentary RNA nucleotides to the 3’ end of the chain
TranscriptionTranscription-DNA guides the production of RNA
Takes place in three phases:•Initiation•Elongation•Termination
•Transcription terminates after a special sequence is transcribed
•Termination sequence (proks-I.e. hairpins)•Polyadenylation sequence (euks)
•The transcript is cut and released
TranscriptionEukaryotic cells modify RNA before it enters the cytoplasm•Both ends of the transcript are processed•Some sections are cleaved and those remaining are spliced together
TranscriptionEukaryotic cells modify RNA before it enters the cytoplasm•Both ends of the transcript are processed•Some sections are cleaved and those remaining are spliced together
•5’ end receives a 5’ cap •3’ end receives a poly-A tail
Protein-coding segment Polyadenylation signal3
3 UTR5 UTR
5
5 Cap Start codon Stop codon Poly-A tail
G P PP AAUAAA AAA AAA…
TranscriptionEukaryotic cells modify RNA before it enters the cytoplasm•Both ends of the transcript are processed•Some sections are cleaved and those remaining are spliced together
Both:•Facilitate the export of the mRNA•Protect mRNA from degradation•Help ribosomes attach
Protein-coding segment Polyadenylation signal3
3 UTR5 UTR
5
5 Cap Start codon Stop codon Poly-A tail
G P PP AAUAAA AAA AAA…
•5’ end receives a 5’ cap •3’ end receives a poly-A tail
TranscriptionEukaryotic cells modify RNA before it enters the cytoplasm•Both ends of the transcript are processed•Some sections are cleaved and those remaining are spliced together
•Introns (non-coding regions) are cut•Exons (coding regions) are spliced together
Pre-mRNA
mRNA
Codingsegment
Introns cut out andexons spliced together
5 Cap
Exon Intron5
1 30 31 104
Exon Intron
105
Exon
146
3Poly-A tail
Poly-A tail5 Cap
5 UTR 3 UTR1 146
TranscriptionQuestion:
What would be the sequence of RNA generated from the following DNA template strand?
DNA: 3’-A T C C G T-5’
TranscriptionQuestion:
What would be the sequence of RNA generated from the following DNA template strand?
DNA: 3’-A T C C G T-5’mRNA:5’-U A G G C A-3’
Overview
DNA mRNA ProteinTranscription Translation
Translation
Proteins are made from polypeptide polymers, which are made from amino acid monomers
Antibody protein Protein from flu virus
Translation
Proteins are made from polypeptide polymers, which are made from amino acid monomers
Antibody protein Protein from flu virus
Translation
Proteins are made from polypeptide polymers, which are made from amino acid monomers
Antibody protein Protein from flu virus
How is the information in RNA transformed into an amino acid?
Translation-The genetic code
• Only four nucleotides in RNA and 20 amino acids (the genetic code is not like Chinese)
• More like English, different permutations of letters build meaningful words– Only 16 two-letter combinations of
nucleotides (42)– 64 possibilities if three-nucleotide
combinations code for an amino acid
Translation-The genetic code
• Only four nucleotides in RNA and 20 amino acids (the genetic code is not like Chinese)
• More like English, different permutations of letters build meaningful words– Only 16 two-letter combinations of
nucleotides (42)– 64 possibilities if three-nucleotide
combinations code for an amino acid
Translation-The genetic codeSecond mRNA base
Fir
st
mR
NA
ba
se
(5 e
nd
of
co
do
n)
Th
ird
mR
NA
ba
se
(3 e
nd
of
co
do
n)
•Genetic instructions for a polypeptide are written in RNA as a series of non-overlapping three-nucleotide “words” (codons)
Translation-The genetic codeSecond mRNA base
Fir
st
mR
NA
ba
se
(5 e
nd
of
co
do
n)
Th
ird
mR
NA
ba
se
(3 e
nd
of
co
do
n)
•Genetic instructions for a polypeptide are written in RNA as a series of non-overlapping three-nucleotide “words” (codons)•Ie. 5’-AAG-3’=lysine
Translation-The genetic codeSecond mRNA base
Fir
st
mR
NA
ba
se
(5 e
nd
of
co
do
n)
Th
ird
mR
NA
ba
se
(3 e
nd
of
co
do
n)
•Genetic instructions for a polypeptide are written in RNA as a series of non-overlapping three-nucleotide “words” (codons)•Ie. 5’-AAG-3’=lysine•61 code for amino acids
Translation-The genetic codeSecond mRNA base
Fir
st
mR
NA
ba
se
(5 e
nd
of
co
do
n)
Th
ird
mR
NA
ba
se
(3 e
nd
of
co
do
n)
•Genetic instructions for a polypeptide are written in RNA as a series of non-overlapping three-nucleotide “words” (codons)•Ie. 5’-AAG-3’=lysine•61 code for amino acids•3 stop codons
Translation-The genetic codeSecond mRNA base
Fir
st
mR
NA
ba
se
(5 e
nd
of
co
do
n)
Th
ird
mR
NA
ba
se
(3 e
nd
of
co
do
n)
•Genetic instructions for a polypeptide are written in RNA as a series of non-overlapping three-nucleotide “words” (codons)•Ie. 5’-AAG-3’=lysine•61 code for amino acids•3 stop codons•AUG=Methionine or start
Translation-The genetic codeSecond mRNA base
Fir
st
mR
NA
ba
se
(5 e
nd
of
co
do
n)
Th
ird
mR
NA
ba
se
(3 e
nd
of
co
do
n)
•Genetic instructions for a polypeptide are written in RNA as a series of non-overlapping three-nucleotide “words” (codons)•Ie. 5’-AAG-3’=lysine•61 code for amino acids•3 stop codons•AUG=Methionine or start•The code is redundant, but not ambiguous
Translation-The genetic codeSecond mRNA base
Fir
st
mR
NA
ba
se
(5 e
nd
of
co
do
n)
Th
ird
mR
NA
ba
se
(3 e
nd
of
co
do
n)
•Genetic instructions for a polypeptide are written in RNA as a series of non-overlapping three-nucleotide “words” (codons)•Ie. 5’-AAG-3’=lysine•61 code for amino acids•3 stop codons•AUG=Methionine or start•The code is redundant, but not ambiguous•This code is nearly universal
Amino acidattachment site
Hydrogenbonds
Anticodon
3
5
TranslationJust as the architects and builders translate the copied blueprints into the city’s infrastructure, tRNA translates mRNA codons into amino acids
tRNA•Consists of a single strand of RNA•Anticodon on one end (can bind with an mRNA codon)•Corresponding amino acid on upper portion
Translation
Three stages:•Initiation•Elongation•Termination
Translation
Three stages:•Initiation•Elongation•Termination
3355U
UA
ACGMet
GTP GDPInitiator
tRNA
mRNA5 3
Start codon
mRNA binding siteSmallribosomalsubunit
5
P site
Translation initiation complex
3
E A
Met
Largeribosomalsubunit
•Initiation begins when mRNA binds to the small sub-unit of a ribosome
Translation
Three stages:•Initiation•Elongation•Termination
3355U
UA
ACGMet
GTP GDPInitiator
tRNA
mRNA5 3
Start codon
mRNA binding siteSmallribosomalsubunit
5
P site
Translation initiation complex
3
E A
Met
Largeribosomalsubunit
•Initiation begins when mRNA binds to the small sub-unit of a ribosome•Charged tRNA binds to the start codon
Translation
Three stages:•Initiation•Elongation•Termination
3355U
UA
ACGMet
GTP GDPInitiator
tRNA
mRNA5 3
Start codon
mRNA binding siteSmallribosomalsubunit
5
P site
Translation initiation complex
3
E A
Met
Largeribosomalsubunit
•Initiation begins when mRNA binds to the small sub-unit of a ribosome•Charged tRNA binds to the start codon•Large ribosomal sub-unit binds•Cluster is called the initiation complex
Translation
Three stages:•Initiation•Elongation•Termination
Amino endof polypeptide
mRNA
5
3E
Psite
Asite
GTP
GDP
E
P A
E
P A
GDPGTP
Ribosome ready fornext aminoacyl tRNA
E
P A
•New tRNAs come in and bind at A site as their complimentary condon is made available
TranslationThree stages:•Initiation•Elongation•Termination
Amino endof polypeptide
mRNA
5
3E
Psite
Asite
GTP
GDP
E
P A
E
P A
GDPGTP
Ribosome ready fornext aminoacyl tRNA
E
P A
•New tRNAs come in and bind at A site as their complimentary condon is made available•New amino acids is bonded to the growing chain
TranslationThree stages:•Initiation•Elongation•Termination
Amino endof polypeptide
mRNA
5
3E
Psite
Asite
GTP
GDP
E
P A
E
P A
GDPGTP
Ribosome ready fornext aminoacyl tRNA
E
P A
•New tRNAs come in and bind at A site as their complimentary condon is made available•New amino acids is bonded to the growing chain•Complex moves to free the A site
TranslationThree stages:•Initiation•Elongation•Termination
Amino endof polypeptide
mRNA
5
3E
Psite
Asite
GTP
GDP
E
P A
E
P A
GDPGTP
Ribosome ready fornext aminoacyl tRNA
E
P A
•New tRNAs come in and bind at A site as their complimentary condon is made available•New amino acids is bonded to the growing chain•Complex moves to free the A site•tRNA that shifted to E site exits
TranslationThree stages:•Initiation•Elongation•Termination
Amino endof polypeptide
mRNA
5
3E
Psite
Asite
GTP
GDP
E
P A
E
P A
GDPGTP
Ribosome ready fornext aminoacyl tRNA
E
P A
•New tRNAs come in and bind at A site as their complimentary condon is made available•New amino acids is bonded to the growing chain•Complex moves to free the A site•tRNA that shifted to E site exits•New tRNA binds at the A site
Translation
Three stages:•Initiation•Elongation•Termination
•Termination occurs when a ribosome encounters a stop codon•Release factors bind •mRNA and polypeptide are released
Releasefactor
3
5Stop codon(UAG, UAA, or UGA)
5
32
Freepolypeptide
2 GDP
GTP
5
3
TranscriptionQuestion:
What polypeptide sequence would be generated from the following DNA template strand?
DNA: 3’-T T C A G T-5’
Second mRNA base
Fir
st
mR
NA
ba
se
(5 e
nd
of
co
do
n)
Th
ird
mR
NA
ba
se
(3 e
nd
of
co
do
n)
TranscriptionQuestion:
What polypeptide sequence would be generated from the following DNA template strand?
DNA: 3’-T T C A G T-5’RNA: 5’-A A G U C A-3’Peptide sequence: Lysine, serine
Second mRNA base
Fir
st
mR
NA
ba
se
(5 e
nd
of
co
do
n)
Th
ird
mR
NA
ba
se
(3 e
nd
of
co
do
n)
Summary
DNA mRNA ProteinTranscription Translation
Point mutations
Impacts of genetic mutation on gene expression
• Point mutations- change in a single base pair– Base-pair substitution
• Silent-no change in polypeptide• Missense (substituted amino acid)• Nonsense (early stop codon)
Point mutations
Impacts of genetic mutation on gene expression
• Point mutations- change in a single base pair– Insertion and deletion
• Addition or loss of base pairs• Causes a frame shift
You should understand:
• The process of gene expression
• The impact of point mutations on gene expression
DNA mRNA ProteinTranscription Translation