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Ch. 10 – From DNA to Protein

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Protein Synthesis

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Metabolism and Gene Expression n  Inheritance of metabolic diseases suggests that genes

coded for enzymes

n  Diseases (phenotypes) caused by non-functional gene product (Tay-sachs, PKU (phenylketonuria), albinism)

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Transcription

(nucleus)

PROTEINS

Translation

(cytoplasm)

The Central Dogma

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RNA vs. DNA

RNA DNA

Single strand

Double strands

Ribose Deoxyribose

Uracil (U) Thymine (T)

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Transcription n  the formation of a

specific RNA sequence from a specific DNA sequence

n  requires: -  a DNA template -  nucleoside

triphosphates (ATP, GTP, CTP, UTP) as substrates

-  RNA polymerase

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§  RNA polymerase binds only to promoter.

§  TATA box §  Transcription factors

(proteins) help RNAP to bind.

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§  RNAP moves along the DNA template from 3’ to 5’

§  RNAP adds complementary nucleotides to the growing mRNA.

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§  RNAP reaches termination site.

§  Transcription stops. §  mRAN falls out.

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Transcription

Transcription Animation http://bcs.whfreeman.com/hillis1e/

#667501__674147__

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Eukaryotic DNA has “junk”

n  Exons – (expressed/coding) segment of DNA

n  Introns – in-between (non-coding) sequence

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RNA splicing enzymes

Not smurfs! “snurps”

n  snRNPs (small nuclear ribonucleoproteins)

n  Spliceosome - snRNPs binds to

consensus sequences

- cut and paste gene

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Alternative RNA splicing n  different segments treated as exons

Defining a gene is getting more complicated!

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RNA Splicing

RNA Splicing http://bcs.whfreeman.com/hillis1e/#667501__674148__

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A A mRNA

5'

3'

G P P P

50-250 A’s

More post-transcriptional processing n  Adding 5ʹ′ GTP cap and poly-A tail: -  protects mRNA on its trip from nucleus to cytoplasm

from (hydrolytic) enzymes in cytoplasm -  facilitates binding of RNAP

n  Neither 5’ cap and poly-A tailget translated into proteins

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Check Your Understanding

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AUGCGUGUAAAUGCAUGCGCC!mRNA

mRNA codes for proteins in triplets

TACGCACATTTACGTACGCGG!DNA

AUGCGUGUAAAUGCAUGCGCC!mRNA

Met Arg Val Asn Ala Cys Ala!protein

?�

codon

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Cracking the Code

Animated Tutorial 10.3 – Deciphering the Code http://bcs.whfreeman.com/hillis1e/

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The Code n  is the code for ALL

life!

n  strongest support for a common origin for all life

n  redundant (several codons for each amino acid) but not ambiguous (code for the different amino acids)

n  3rd base (3’ end) “wobble”

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Transfer RNA structure n  “Clover leaf” structure n  anticodon on “clover leaf” end, amino acid

attached on 3ʹ′ end

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Charging tRNA n  Aminoacyl-

tRNA synthetase bonds amino acid to tRNA

n  Energy is required

n  ATP → AMP

n  bond is unstable so it can release amino acid at ribosome easily

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Ribosomes n  A site (aminoacyl-tRNA) holds tRNA carrying next

amino acid to be added to chain

n  P site (peptidyl-tRNA) holds tRNA carrying growing polypeptide chain

n  E site (exit) empty tRNA leaves ribosome from exit site

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Building a polypeptide n  Initiation brings together

mRNA, ribosome subunits, initiator tRNA

n  Elongation adds amino acids based on codon sequence

n  Termination adds end codon

1 2 3

Leu

Leu Leu Leu

tRNA

Met Met Met Met

P E A mRNA 5' 5' 5' 5'

3' 3' 3' 3' U U A A A A C

C C

A U U G G G U U

A A A A C

C C

A U U G G G U U

A A A A C

C C

A U U G G G U U A A A C

C A U U G G

Val Ser Ala Trp

release factor

A A A

C C U U G G 3'

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Protein Synthesis

Animated Tutorial 10.4 – Protein Synthesis http://bcs.whfreeman.com/hillis1e/

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Initiation

n  small ribosomal subunit binds to mRNA recognition sequence

n  Methione-charged tRNA binds to AUG.

n  Large subunit joins

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Elongation n  Second tRNA enters A

site n  Bond breaks between

tRNA in P site and its amino acid

n  peptide bond forms between that amino acid and the amino acid on tRNA in A site

n  First tRNA moves to E site, dissociates from complex

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Elongation

n  Elongation occurs as the steps are repeated, assisted by elongation factors.

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Elongation

n  a stop codon enters A site

n  Stop codon binds a protein release factor

n  Polypeptide chain separates from the ribosome

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Protein Targeting

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Protein Modifications

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Check Your Understanding

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Check Your Understanding

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Can you tell the story?

DNA

pre-mRNA

ribosome

tRNA

amino acids

polypeptide

mature mRNA

5' GTP cap

poly-A tail

large ribosomal subunit

small ribosomal subunit

aminoacyl tRNA synthetase

E P A

5'

3'

RNA polymerase

exon intron

tRNA

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Protein Synthesis in Prokaryotes

Bacterial chromosome

mRNA

Cell wall

Cell membrane

Transcription

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Prokaryote vs. Eukaryote genes n  Prokaryotes - DNA in cytoplasm - circular chromosome - naked DNA - no introns

n  Eukaryotes - DNA in nucleus -  linear chromosomes - DNA wound on

histone proteins -  introns vs. exons

eukaryotic DNA

exon = coding (expressed) sequence

intron = noncoding (inbetween) sequence

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n  Transcription and translation are simultaneous in bacteria

n  DNA is in cytoplasm

n  no mRNA editing

n  ribosomes read mRNA as it is being transcribed

Translation in Prokaryotes

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Translation in Eukaryotes