Offner Protein Synthesis Flashcards
Factors required for protein synthesis
IF2-GTP
EFTU-GTP
EF-G-GTP
RF-3-GTP
- add “e” for eukaryotic
How are amino acids attached to tRNAs?
Uses synthetase for both steps
- amino acids activated with ATP to aminoacyl adenylate
- tRNA added to form aminoacyl-tRNA + AMP
Requires cleavage of 2 high-energy bonds
Structure of amino acid synthetase
- activation site
- hydrolytic site (corrects errors)
- aa-tRNA that leave enzyme cannot be error corrected
Initiation of prokaryotic protein synthesis
Initiation complex binds to Shine-delgarno sequence
- initiator tRNA binds in P site
Prokaryotic protein initiation complex
Contains tRNA, fMET, IF2-GTP
- 30S and 50S bind, causing release of IF1, IF3, IF2-GDP
Which subunit and part binds with the Shine-Dalgarno sequence
It base pairs with 3’ end of 16S rRNA of the 30S subunit
How is fMET created?
Formyl group added to initiator tRNA after activation with Methionine
Translation elongation mechanism
- EF-Tu-GTP puts tRNA in A site
- Peptidyl transferase creates peptide bond
- EF-G-GTP cases translocation of ribosome
Translation Termination mechanism
- RF1/2 bind A site, RF3-GTP binds common site
- Peptidyl transferase cleaves ester bond via GTP
- release of everything
How is peptidyl transferase made, and which subunit is a part of?
Ribozyme, part of 23S or 28S rRNA
How does EF-G-GTP cause translocation of the ribosome
Hydrolysis of GTP
What is the common site on the prokaryotic ribosome
Where TF, EF and RF all bind
- ensures stepwise progression of synthesis (not all at same time)
What do polyribosomes do
Make multiple proteins from the same message
- simultaneous message transcription
What do Tetracycline, Chloramphenicol and Puromycin do, and do they affect prokaryotes or eukaryotes?
Inhibit translation, all affect prokaryotes, only puromycin affects eukaryotes
Tetracyline
Prokaryotes only
- blocks binding of aminoacyl-tRNA to A site
Chloroamphenicol
Prokaryotes only
- resembles peptide bond
- inhibits peptidy transferase
Puromycin
Prok and Euk
- enters A site
- accepts polypeptide chain
- blocks translocation
How many ATP/GTP hydrolyzed per 100 amino acids for:
Activation, Initiation, Elongation EF-Tu and EF-G, Termination
Activation: 200ATP
Initiation: 1GTP
Elongation EF-TU: 99 GTP
Elongation EF-G: 99GTP
Termination RF-3: 1GTP
400 total
Eukaryotic translation initiation
- small subunit binds at cap
- 40S scans from cap to find Kozak sequence (CCACCAUGG)
- GTP used to join 60S to 40S
Cap binding proteins
eIF4G - scaffold
eIF4A- Helicase
eIF4B - ATPase
eIF4E - binds eIF3 on 40S
Role of eIF4B and eIF4A
Combine to unwind secondary structure
Role of eIF4E
Facilitates binding of 40S and initiator tRNA-eIF2-GTP complex
Role of eIF5
Releases 40S associated factors, combines it with 60S using GTP to begin translation
How are ribosomes recycled in eukaryotes
mRNA loop structure
HCV lifecycle
1) tRNA into cell
2) creation of viral proteins
3) negative RNA produced, replicated to create positive RNA for export
HCV 5A and 5B proteins
5A: replication complex protein
5B: RNA-dependent RNA Pol
How is HCV treated
Target 5A and 5B
Is HCV RNA capped?
No, but it contains IRES: structural feature which allows ribosome to bind for replication
Does the Shine-Dalgarno sequence contain mainly pyrimidines or purines
‘tis purine-rich
