From gene to protein. Our plan: Overview gene expression Walk through the process –Review...

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