Protein Synthesis 1 Flashcards
What are the three phases of translation?
Initiation
Elongation
Termination
What is the role of IF-1 and IF-3 in prokaryotic initiation?
Bind to the 30s (small) ribosomal subunit
What happens in prokaryotic initiation after the formation of IF-1/IF-3/30s complex?
IF-2GTP and N-formylmethionyl tRNA bind the complex and join to mRNA
What stimulates the binding of the 50s subunit?
- Release of IF1 and IF3
- Hydrolysation of GTP to GDP and then release of IF2
What two processes happen simultaneously during eukaryotic translation initiation?
43s preinitiation complex formation
mRNA preparation
How is the 43s preinitiation complex formed?
- eIF1,1A,and 3 bind the 40s ribosomal subunit
- methionyl-tRNA, eIF2-GTP, and eIF5 bind ribosome complex
What happens during mRNA preparation?
- PABP binds the 3’ poly-A tail
- eIF-4F complex binds the 5’ end of the mRNA
What is the function of eIF4G?
Links the eIF4F complex to PABP
What is the function of eIF4E?
-binds the 5’ m7 G cap of mRNA
What is the function of eIF4B?
-unwinds secondary structures in the mRNA
What is the function of eIF3?
Binds both the mRNA and preinitiation complex, linking the two structures.
This new 48s initiation complex starts scanning.
What is the role of the kozak sequence?
Identifies the correct AUG start codon
usually the most 5’ AUG sequence
What is scanning?
The 48s initiation complex utilizes ATP in order to scan the mRNA for the start codon (AUG)
What occurs when the 48s initiation complex reaches the correct start codon?
eIF5 triggers eIF2GTP–>GDP, causing the eIFs to lose affinity for the complex and dissociate.
What is the role of eIF5B?
Has a bound GTP and recruits the 60s large ribosomal subunit to form the 80s ribosome via hydrolysis of GTP
Why are there two separate tRNAs that code for Met?
One for the initiation Met
One for internal Mets
What are the 3 tRNA binding sites in ribosomes and their function?
A (aminoacyl or acceptor)- holds incoming tRNA
P (peptidyl) - carries growing polypeptide
E (exit) - empty tRNA exits
What is the function of eEF1a-GTP?
- Bring the charged tRNA to the ribosome
- conformational change by the binding of the tRNA induces GTP hydrolysis, releasing eIF1a-GDP and leaving the tRNA in the A site.
What is responsible for catalyzing the formation of new peptide bonds?
-Peptidyltransferase of the 60s ribosomal unit
What is the function of eEF-2GTP?
mediates the translocation of the ribosome down the mRNA
How does diptheria toxin work?
Inactivates eEF2, preventing elongation
How does ricin work?
Cleaves adenine residue from the 60s subunit, resulting in the inability of eEF2 to bind
What is the role of eEF1-By?
Exchanges GTP for bound GDP in eEF1a, recharging it and allowing it to bind to another tRNA
What are the three stop codons?
UAA
UAG
UGA
What is responsible for terminating protein synthesis?
Release factors
What is the role of eRF-1?
Recognizes stop codons and binds eRF-3/GTP and enters A site
What results from the binding of eRF1/eRF3 in the A site?
Polypeptide is released from the ribosome
Dissociation of the ribosomal subunits
What are four ways of regulating protein translation?
- Proteins that bind to either the 3’ or 5’ UTRs
- Noncoding RNAs e.g. microRNAs and siRNAs
- Phosphorylation
- Modulation as a result of response to stress, nutrient availability, and growth factor stimulation
How is the translation of ferritin regulated?
When iron is absent, iron regulatory protien binds to the iron response element in the 5’ UTR, blocking translation
What is an example of translation regulation by binding of protein to the 3’ UTR?
Xlerk mRNA localization in Xenopus oocytes. 3’ binding proteins direct the mRNA to specific regions.
How is eIF2 regulated by phosphorylation?
Phorphorylation of eIF-2 and eIF2B by regulatory protein kinases blocks the exchange of bound GDP for GTP.
What is the role of 4E-BP?
Sequesters eIF-4E
When 4E-BP is phosphorylated it releases eIF4E so that the eIF-4F complex can form
What are the roles of ATP and GTP during initiation?
- ATP hydrolysis powers scanning
- eIF-2 undergoes GTP hydrolysis to dissociate the pre-initiation complex
- eIF-5B uses GTP hydrolysis to recruit the large subunit
What are the roles of ATP and GTP during elongation?
- Each tRNA used 1 ATP to charge the tRNA
- eEF-1a uses GTP hydrolysis to bring a charged tRNA to the A site
- eEF-2 uses GTP hydrolysis to power the translocation of the ribosome
What is the function of streptomycin?
Inhibits initiation by binding to and distorting the structure of the 30s ribosomal unit
What is the function of Tetracyclines?
Interacts with the 30s ribosome subunit to block access to the A site and inhibit elongation
What is the function of Chloramphenicol?
Binds and inhibits the peptidyltransferase activity of the 50s ribosome, blocking elongation
What is the function of erythromycin and clindamycin?
inhibit translocation of the ribosome by binding the 50s subunit, blocking elongation
What is the function of Puromycin?
Enters the A site and is incorporated into the peptide, causing premature release of the polypeptide
Inhibits both prokaryotic and eukaryotic protein synthesis
