Trans-translation and ribosomal peptidyl transferase Flashcards
Bacteria face a major problem during trans-translation when ribosomes..
Become stranded on non-stop mRNA
What is non-stop mRNA?
Broken mRNA lacking a stop codon
Can you get multiple ribosomes trapped on non-stop mRNAs?
Yes, in polyribosomes
What is tmRNA used for?
To release incomplete polypeptides (with C-terminal tags) from ribosomes on non-stop mRNA
What does tmRNA resemble?
tRNA lacking an anticodon stem-loop but with an ‘extra’ mRNA-like region (MLR) containing an open reading frame that can be translated
What is the ORF in the tmRNA used for?
To put a C-terminal tag on truncated peptide during release from non-stop mRNA
What gene encodes the 363 nucleotide E. coli tmRNA?
ssrA gene
What tmRNA tag is the target for proteases such as ClpXP?
ANDENYALAA
What does tmRNA (ssrA RNA) work together with?
SmpB protein
Forms quaternary complex with EF-Tu + GTP and binds into ribosomal A site
Outline the steps that follow when tmRNA-SmpB binds in A site?
- Peptide bond formation
- Translocation brings tmRNA ORF into A site
- ORF of tmRNA is now being translated; aatRNA binds into A site
- Translation of tmRNA ORF generates C-terminal tag on truncated protein; peptide bond formation
How many amino acids does the tmRNA ORF encode?
10 amino acid sequence
Are ribosomes recycled?
Yes
Is the alanine in the tag coded for?
No, it’s inserted into empty decoding site
What terminates the synthesis of the tag?
UAA stop codon
What does PTC stand for?
(Ribosomal) peptidyl transferase centre
What are the CCA-3’ ends of tRNAs in the PTC positioned for?
Peptide bond formation by specific H-bonding to P-loop and A-loops of 23S rRNA
What does puromycin resemble?
The 3’ end of aa-tRNA
What is puromycin?
An acceptor substrate for peptidyl transferase
What happens when the nascent peptide is joined to puromycin?
The peptidyl-puromycin falls off ribosome
Is the PTC on the larger or smaller ribosomal unit?
The larger one
What do the donor substrates in the P site need?
2’-OH
What is the 2’-OH of the P site involved in?
Proton shuttle
What are the 3’-CCA ends of tRNAs held in position at the PTC by?
The P-loop and A-loop of 23S rRNA
