Module 5: Transcription & Translation

Question 1

Messenger RNAs (mRNA)

Answer 1

encode for polypeptides

made in the 5’->3’ orientation

Question 2

Transfer RNAs (tRNAs)

Answer 2

read the mRNA code and transfer the appropriate amino acid to a growing polypeptide chain in a process called translation

Question 3

Ribosomal RNAs (rRNAs)

Answer 3

ribosomes composed of rRNA and proteins

translate the RNA message into proteins

Question 4

Promoter

Answer 4

sequence of DNA to which RNA polymerase binds and initiates transcription

Question 5

Consensus Sequence

Answer 5

refers to certain nucleotides that particularly common at each position

Question 6

-10 region

Answer 6

5’ TATAAT 3’

Question 7

-35 region

Answer 7

5’ TTGACA 3’

Question 8

UP (upstream promoter) recognition element

Answer 8

A/T rich

occurs between -40 and -60 positions in promoters that have certain highly expressed genes

Question 9

Main similarities between transcription and DNA replication

Answer 9

• Same fundamental chemical mechanism (creation of phosphodiester bond)
• Same direction of synthesis (5’–>3’)
• Same 3 phases: initiation, elongation, termination

Question 10

Main differences between transcription and DNA replication

Answer 10

• Transcription does NOT require a primer
• Transcription utilizes limited segments of the DNA molecule
• Transcription uses only one of the two DNA strands as a template

Question 11

RNA polymerase

Answer 11

holoenzyme containing 5 subunits
6th subunit: sigma subunit -binds transiently to the core RNA holoenzyme and directs the polymerase to specific binding sites on the DNA
does not have a 3’->5’ proofreading exonuclease activity

Question 12

Sense strand

Answer 12

5’ -> 3’ strand

not used as template

Question 13

Antisense

Answer 13

3’->5’ strand

template strand for RNA polymerase

Question 14

Transcription Initiation

Answer 14

RNA pol core binds to DNA promote -creates closed complex
most predominant sigma subunit: sigma 70
Transcription bubbles forms -open complex (12-15 bp unwounded)

Question 15

Transcription Elongation

Answer 15

sigma 70 dissociates

replaced by the protein NusA -facilitates transcription termination

Question 16

Transcription Termination: Rho factor-independent termination

Answer 16

• Hairpin: region that produces an RNA transcript with self-complementary sequences which folds in on itself (15-20 nucleotides before RNA strand ends)
• Downstream run of 3-8 adenine residues

When the RNA polymerase arrives at this hairpin structure it pauses transcription

Question 17

Transcription Termination: Rho factor dependent termination

Answer 17

requires Rho (ATP dependent RNA-DNA helicase) binds to the 3’ end of the nascent transcript (CA-rich site=rut)

Rho proceeds down the transcript towards the 3’ end unwinding the 3’ end of the transcript, RNA pol and pr- factors released

Question 18

mRNA transcript complements…

Answer 18

antisense strand

same sequence as sense strand (except U)

Question 19

Genetic Code

Answer 19

61/64=sense codons (code for aa)
other 3=nonsense codon (stop codons)
degenerate/redundant: most aa have more than 1 codon
unambiguous: each specific trinucleotide encodes only 1 aa

Question 20

Wobble Hypothesis

Answer 20

most aa in table have same 2 codon letters but differ in third codon letter

Question 21

Shine-Dalgarno sequence

Answer 21

consensus sequence allows for proper positioning of the start codon (AUG) on the mRNA relative to the ribosome, which allows for initiation of translation

Question 22

tRNA structure

Answer 22

bottom: anticodon triplet (complementary to mRNA codon)
top: acceptor stem (aa attachment site)

Question 23

Aminoacyl-tRNA synthetases (aaRS)

Answer 23

enzymes whose function is to covalently attach amino acids to their respective tRNA molecules

occurs between the carboxylate of the amino acid and the ribose 3’ OH of the invariant 3’ terminal adenosine residue on the tRNA

Question 24

70S ribosome

Answer 24

• 50S and 30S subunit
• 3 tRNA binding sites: E, P, A
• binds and decodes mRNA on 30S subunit
• Binds tRNA -fills gaps b/w 50S and 30S subunits
• allows newly synthesized chain to exit from 50S (tunnel)
• allows peptidyltransferase runs to occur on 50S which covalently attaches aa through peptide bonds

Question 25

Step 1: Activation of aa

Translation

Answer 25

carboxyl group of each amino acid is “activated” to facilitate formation of the peptide bond
aminoacyl-tRNA synthetases -prep or “charge” each tRNA ready for protein synthesis

Question 26

Step 2: Initiation

Translation

Answer 26

• AUG binds tRNA, fMet
• 30S binds two initiation factors, IF1 and IF3
• IF3 prevent the 30S and 50S subunits from interacting prematurely
• mRNA bind to Shine-Dalgarno Sequence, allowing binding of IF2-GTP (recruit fMet-tRNA)
• 50S assembled
• GTP hydrolyzed to GDP+Pi -all IFs dissociate

Question 27

Step 3: Elongation

Translation

Answer 27

• first tRNA bound to P site of 50S
• second enters A site (allowed through GTP hydrolysis)
• peptide bond formed, transfer aa moved to A site, P site tRNA moves to E site (released into cytosol)
• A tRNA moves to P site

Question 28

Step 4: Termination

Translation

Answer 28

3 codons: UAA, UAG, UGA

• Once in A site, release factor (RF) bind to A site on 50S
• leads to the subsequent hydrolysis of the ester linkage between the nascent polypeptide and the tRNA in the P-site and final release of the completed polypeptide
• 30S/50S subunits dissociates and release of all associated factors

Question 29

3’ end

Answer 29

poly A tail

Question 30

5’ end

Answer 30

methylated cap