SURVEY OF BIOCHEMISTRY Transcription and Translation 1 PRS

SURVEY OF BIOCHEMISTRY Transcription and Translation 1

PRS Which of the following is not an example of a DNA-binding motif in proteins? 1. Beta Sheet 2. Helix-turn-Helix 3. Zinc Finger 4. Leu Zipper 2

PRS • Which amino acid contains a hydroxyl in its side chain? 1. Threonine 2. Methionine 3. Aspartic Acid 4. Phenylalanine 3

Transcription is the process by which RNA is produced from a DNA template. Steps involved in transcription § Initiation § Elongation § Termination 4

Transcription DNA has to separate at the position where transcription occurs. Fig. 26 -3 5

Transcription Sense Strand Newly synthesized RNA has the same general sequence as the sense strand; U in RNA, T in DNA 5’-…GATCCGTAGGTC…-3’ 3’-…CTAGGCATCCAG…-5’ Antisense Strand (template) 5’-…GATCCGTAGGTC…-3’ 5’----GAUCCGUAGGUC… 3’ 3’-…CTAGGCATCCAG…-5’ 6

Transcription Initiates at Promoter Sequences What is a gene? Example of E. coli Lactose Operon Organization Fig. 26 -4 P: Promoter O: Operator I: Inhibitor 7

Structural genes follow promoter sequences In E. coli, the lactose operon consists of 3 structural genes: § Beta-Galactosidase (Z) § Galactoside Permease (Y) § Thiogalactoside Transacetylase (A) 8

E. coli Promoter Sequences Notice how sequence is numbered! Usually single purine 9

Prokaryotic RNA Polymerases Core enzyme Holoenzyme The sigma factor binds specifically to the promoter. Core enzyme affinity to DNA ~5 x 10 -12 M. = sigma factor 10

Elongation: RNA Chain Growth 3’-OH group acts as a nucleophile, attacking the alpha phosphate group, with subsequent release of inorganic pyrophosphate. 11

DNA Supercoiling Negative supercoiling Positive supercoiling 1. RNA polymerase is processive 2. Once an RNAP has moved from the promoter, another can bind 3. Transcription is fast: constitutive enzymes vs. inducible enzymes 12

Termination of Transcription Because the newly synthesized RNA is single-stranded, hairpin structures can easily form from the termination sequence. 13

Hairpin Structure in RNA G-C rich bp region What happens when a known terminator sequence is missing? Poly-U tail 14

Termination: Rho Factor Helicase: RNA-RNA and RNA-DNA complex unwinding. 15

Transcription in Eukaryotes • RNA Polymerase I – Precursors to r. RNA • RNA Polymerase II – Precursors to m. RNA • RNA Polymerase III – Precursors to 5 S r. RNA, t. RNA – Precursors to other small RNA’s 16

Eukaryotic Transcription Factors Transcription factors are req’d to initiate transcription, and facilitate elongation in eukaryotes. Eukaryotic Termination: Not well understood 17

Portion of the Genetic Code Wobble 18

Reading Frames 19

Translation: Protein Synthesis • Messenger RNA – Template • Transfer RNA – Carries amino acids • Ribosomal RNA – Aids protein synthesis 20

t. RNA Structures 2 D 3 D 21

t. RNA’s have modified bases Be able to name, draw, recognize structures from Figure 27 -4. 22

Aminoacyl t. RNA Synthetases Aminoacyl t. RNA synthetases add aa’s to t. RNA. Step 1. 23

Aminoacyl t. RNA Synthetases Step 2. + t. RNA 24

Ribosome 25

Sites in the Ribosome A Aminoacyl E P A P Peptidyl E Exit 26

Chain Initiation in Translation Start Codon: AUG Methionine 27

Shine-Dalgarno Sequence upstream downstream 28

Initiation 29

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Translation: Polypeptide Synthesis Peptidyl Transferase Activity 31

Elongation 32

Administrative Updates • Exam 1 on Thursday, 4 -5 pm – 20 questions – Short answers and Multiple Choice – Closed-book – Calculators allowed – No cell phones, blackberries, etc. • Office Hours: M, T, R 2 -4 pm 33