molecular bio exam #4!!! last half

Depdendent on ATP or GTP for energy:

Aminoaccyl bond formation, translocation, ribosome release

Not depdentedent on ATP or GTP for energy

peptide bond formation

would insertion of 6 nucleotides, deletion of 3 change reading frame

no

If threonine aminoacyl-tRNa synthetase confronted w both serine and threonine AA. What would happen

Ser will bind to acetylation site and proofreading site

Ski7 recruitment in response ot transcription of extended poly-lysine sequence initaties

non-stop decay

Function of Shine-Dalgarno sequence

Helps mRNA bind to small subunit, align start codon to p site

Hydrolysis of GTP to GDP help facilitate

IF-1, IF-2, IF-3 leave

What other eukaryotic factor heps fufill diff part of function of Shine dlagarno secquence

eIF4F

What factor is not bound to GTP or ATP or involved in its hydrolysis

eIF3

what is a feature of eukaryotic translation initation that is different from bacteria

mRNA binding occurs after initation tRNA binding

eIF4G function

tethers mRNA to small subunit, tethers 5’ cap of mRNA to poly A tail

which process in eukaryotes ensures that first AUG encountered after kozack sequence is the starting codon

scanning

If you deleted gene coding for EF-Ts, cells lacking this protein would lack energy for

accomidation

accomidation

animoacyl-tRNA fullly enters A site of ribosome

What are the two steps of tRNA activation?

1. Adenylylation 2. tRNA charging

ribosome recycling factor, rrf

Drives ribosomal subunits apart

RF-2

dives last peptidyl-transferase

RF-1

recognizes stop codon UAA and UAG

RF-3

GTP hydrolysis to detach release factors

EF-G

provides energy for release of tRNA from ribosome, moving it through it too

EF-Tu

binds to aminoacyl-tRNA, delivers to the A site

EF-Tu mechanism

requires GTP bind TRNA, hydrolzed, allows tRNA to be added to ribosome

EF-Ts

exchanging GDP for GTP on EF-Tu, recycling EF-Tu back to GTP bound state

EF-P

Peptide bond formation at start of translation

How do Aminoacyl-tRNA synthetases recognize tRNAs

identity nuleotides

What is adenylylation?

AMP binds to AA

What happens during tRNA charging?

specific AA attached to tRNA, energy from ATP, creates charged tRNA

Where is the aminoacyl group transferred?

From AMP to the CAA arm of the tRNA.

How many aminoacyl-tRNA synthetases exist?

20, one for each amino acid.

What does class 1 synthetase do?

Attaches the AA to the 2' OH of CCA.

What does class 2 synthetase do?

Attaches the AA to the 3' OH of CCA.

What ensures the accuracy of the genetic code?

specificity and proofresding of aminoacyl-tRNA synthetases.

What is the second genetic code?

The interaction between synthetases and different tRNAs.

Why do synthetases need proofreading?

Because many amino acids, like Valine and Isoleucine, are structurally similar.

Why is Valine problematic for Ile-tRNAᶦˡᵉ synthetase?

Valine is small enough to fit in the isoleucine binding site.

What happens if a large AA tries to attach to a smaller AA’s tRNA?

It doesn’t fit into the binding site of the synthetase.

Can Ile fit into Val-tRNA synthetase?

No, it's too large to fit.

Does Val-tRNA synthetase have a proofreading site?

No

Can Valine fit in the acylation site of Ile-tRNAᶦˡᵉ synthetase?

Yes

How does Ile-tRNAᶦˡᵉ synthetase fix Valine mispairing?

The proofreading site detects the small Val-tRNAᶦˡᵉ and removes it.

What happens when an incorrect AA fits into the proofreading site?

The AA-tRNA is hydrolyzed and the process restarts.

Why does proofreading work in this case?

Only smaller incorrect amino acids fit into the proofreading site.

How does Ile-tRNAᶦˡᵉ synthetase ensure correct AA binding?

acylation site and a proofreading site to detect incorrect small AAs.

What happens if the wrong AA is transferred to a tRNA?

It is hydrolyzed and removed before the AA-tRNA is released.

What is the initiator tRNA in prokaryotes?

tRNAᶠᴹᵉᵗ (formyl-Met)

What is the internal methionine tRNA in prokaryotes?

tRNAᴹᵉᵗ

How many Met-tRNA synthetases do prokaryotes have?

Only one, it recognizes both tRNAs.

What does transformylase do in prokaryotes?

Converts Met into N-formylmethionine (fMet).

Is the formyl group added internally?

No, only added for the first AA.

What is the initiator tRNA in eukaryotes?

tRNAⁱᴹᵉᵗ

What is the internal methionine tRNA in eukaryotes?

tRNAᴹᵉᵗ

Is methionine modified in eukaryotes?

no

Which tRNA is recognized by the ribosome initiation complex in eukaryotes?

Only tRNAⁱᴹᵉᵗ.

Why do many mature proteins lack Met at the start?

Aminopeptidases remove N-terminal Met during processing.

What are the steps of translation?

1. Processed mRNA exits nucleus 2. Ribosome assembles on mRNA 3. tRNA binds codon 4. Peptide bond forms 5. Ribosome shifts 3 nucleotides.

What moves along the mRNA?

The ribosome, 3 nucleotides at a time.

What did Harry Noller’s experiment show?

Ribosome's catalytic activity comes from rRNA, not protein.

What does puromycin do?

Mimics tRNA, enters A site, causes premature termination.

Why is puromycin a strong antibiotic?

causes translation to stop by blocking peptide bond formation.

What is a ribozyme?

RNA with enzymatic activity.

What are the ribosome tRNA binding sites?

A site (entry), P site (peptide bond), E site (exit).

What are the 4 stages of translation?

Activation, initiation, elongation, termination.

What happens during initiation?

mRNA and aminoacyl-tRNA bind to small subunit.

What happens during elongation?

tRNA binding, peptide bond formation until stop codon.

What happens during termination?

Translation ends, ribosome dissociates, protein released.

Is protein folding part of translation?

Yes, during termination.

What is the final ribosome structure?

Small + large subunit + mRNA + initiator tRNA.

What is the Shine-Dalgarno sequence?

Helps mRNA bind to small subunit, and Helps align start codon to P site

What is the eukaryotic equivalent of Shine-Dalgarno?

The Kozak sequence.

What does IF-1 do in bacteria?

Blocks the A site to prevent premature tRNA binding.

What does IF-2 do in bacteria?

Escorts initiator tRNA to the ribosome.

What does IF-3 do in bacteria?

Prevents the large subunit from binding prematurely.

What powers the formation of the bacterial initiation complex?

GTP hydrolysis.

What happens in Step 1a of bacterial initiation?

IF-1 and IF-3 bind the 30S small subunit.

What happens in Step 1b? bacterial

mRNA is recruited using Shine-Dalgarno to position start codon.

What happens in Step 2? bacterial

IF-2 binds initiator tRNA (fMet-tRNAᶠᴹᵉᵗ) and places it at P site.

Why can only fMet-tRNAᶠᴹᵉᵗ bind the P site? bacterial

Because of a unique sequence; other tRNAs can't bind there.

What happens in Step 3? bacterial

50S binds, IFs are released, and initiation complex is complete.

What are the 3 basic steps of bacterial initiation?

1. Blocking sites, 2. Loading initiator tRNA, 3. Loading large subunit.

Does mRNA attach during step 1 in eukaryotes?

No, unlike bacteria.

What does eIF2 do?

Binds initiator tRNA (tRNAⁱᴹᵉᵗ) to ribosome

Is eIF2 GTP-bound?

Yes, like IF-2.

eIF5B

GTPase that joins the initiator tRNA to the P site.

Does eukaryotic initiator tRNA need the start codon to load?

No, unlike in bacteria.

What does eIF4F complex do?

Binds the 5' cap and escorts mRNA to the small subunit.

What are the parts of eIF4F?

eIF4E (cap binding), eIF4A (ATPase/helicase), eIF4G (adapter).

What is required for eIF4F/mRNA complex binding?

Hydrolysis of ATP by eIF4A.

What happens during scanning?

Pre-initiation complex moves along mRNA to find start codon.

What identifies the start codon in eukaryotes?

Kozak sequence, similar to Shine-Dalgarno.

How many GTPs and ATP are required in eukaryotic large subunit loading?

2 GTPs (eIF2, eIF5B) and 1 ATP (eIF4A).

What are bacterial polysomes?

Multiple ribosomes translating same mRNA while being transcribed.

What allows polysome formation in eukaryotes?

eIF4G binding poly-A binding protein to circularize mRNA.

What is IRES? Internal Ribosome Entry Site

a way to recognize mRNA without 5’ cap

Why is IRES useful in research?

Allows expression of 2 genes from one mRNA transcript.