Release of nonstop ribosomes is essential.

mBio (2014-11-13)
Heather A Feaga, Patrick H Viollier, Kenneth C Keiler
ABSTRACT

Bacterial ribosomes frequently translate to the 3' end of an mRNA without terminating at a stop codon. Almost all bacteria use the transfer-messenger RNA (tmRNA)-based trans-translation pathway to release these "nonstop" ribosomes and maintain protein synthesis capacity. trans-translation is essential in some species, but in others, such as Caulobacter crescentus, trans-translation can be inactivated. To determine why trans-translation is dispensable in C. crescentus, a Tn-seq screen was used to identify genes that specifically alter growth in cells lacking ssrA, the gene encoding tmRNA. One of these genes, CC1214, was essential in ΔssrA cells. Purified CC1214 protein could release nonstop ribosomes in vitro. CC1214 is a homolog of the Escherichia coli ArfB protein, and using the CC1214 sequence, ArfB homologs were identified in the majority of bacterial phyla. Most species in which ssrA has been deleted contain an ArfB homolog, suggesting that release of nonstop ribosomes may be essential in most or all bacteria. Genes that are conserved across large phylogenetic distances are expected to confer a selective advantage. The genes required for trans-translation, ssrA and smpB, have been found in >99% of sequenced bacterial genomes, suggesting that they are broadly important. However, these genes can be deleted in some species without loss of viability. The identification and characterization of C. crescentus ArfB reveals why trans-translation is not essential in C. crescentus and suggests that many other bacteria are likely to use ArfB to survive when trans-translation is compromised.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
D-(+)-Xylose, ≥99%
Sigma-Aldrich
D-(+)-Xylose, ≥99%
Sigma-Aldrich
D-(+)-Xylose, BioXtra, ≥99%
Sigma-Aldrich
D-(+)-Xylose, BioUltra, ≥99.0% (sum of enantiomers, HPLC)
Sigma-Aldrich
Nitrilotriacetic acid trisodium salt, Sigma Grade, ≥98%
Sigma-Aldrich
Nitrilotriacetic acid disodium salt, Sigma Grade, ≥99%
Xylose, European Pharmacopoeia (EP) Reference Standard