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SsrA‐mediated peptide tagging caused by rare codons and tRNA scarcity
Author(s) -
Roche Eric D.,
Sauer Robert T.
Publication year - 1999
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/18.16.4579
Subject(s) - biology , transfer rna , messenger rna , stop codon , release factor , ribosome , translation (biology) , protein biosynthesis , rna , peptide , genetics , microbiology and biotechnology , biochemistry , gene
SsrA RNA mediates the addition of a C‐terminal peptide tag (AANDENYAL AA ) to bacterial proteins translated from mRNAs without in‐frame stop codons. This process involves both tRNA‐ and mRNA‐like functions of SsrA and targets the tagged proteins for degradation. By designing an SsrA variant that adds a peptide tag (AANDENYAL DD ) that does not result in rapid degradation, we show that tagging of a model protein synthesized from an mRNA without stop codons can be detected both in vivo and in vitro . We also use this assay to demonstrate that ribosome stalling at clusters of rare arginine codons in mRNA is sufficient to recruit and activate the SsrA peptide tagging system. An essential requirement for tagging at rare AGA codons is a scarcity of the cognate tRNA; supplemental tRNA AGA suppresses tagging, and depleting the available pool of tRNA AGA enhances tagging and reveals tagging caused by single rare AGA codons. Protein tagging at sites corresponding to rare codons appears to involve SsrA action at an internal mRNA site rather than at the 3′ end of a cleaved mRNA.