Mol Cell. 2009 Apr 24;34(2):201-11.

The plasticity of a translation arrest motif yields insights into nascent polypeptide recognition inside the ribosome tunnel.

Source

Genetics and Biochemistry Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

Abstract

The recognition of a C-terminal motif in E. coli SecM ((150)FXXXXWIXXXXGIRAGP(166)) inside the ribosome tunnel causes translation arrest, but the mechanism of recognition is unknown. Whereas single mutations in this motif impair recognition, we demonstrate that new arrest-inducing peptides can be created through remodeling of the SecM C terminus. We found that R163 is indispensable but that flanking residues that vary in number and position play an important secondary role in translation arrest. The observation that individual SecM variants showed a distinct pattern of crosslinking to ribosomal proteins suggests that each peptide adopts a unique conformation inside the tunnel. Based on the results, we propose that translation arrest occurs when the peptide conformation specified by flanking residues moves R163 into a precise intratunnel location. Our data indicate that translation arrest results from extensive communication between SecM and the tunnel and help to explain the striking diversity of arrest-inducing peptides found throughout nature.

PMID:: 19394297; [PubMed - indexed for MEDLINE]
PMCID:: PMC2704006

Free PMC Article

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Fig. 1

Isolation of second-site mutations that suppress a defect in SecM-mediated translation arrest

(A) A plasmid encoding the translation arrest-deficient Q160P allele of SecM(Eco) was randomly mutagenized, and intragenic mutations that suppress the defect were isolated in a LacZα reporter screen. Positions where the sequence of the suppressor mutants (Sup1–6) and wild-type SecM differ are indicated. SecM(Eco) residues that were previously shown to be required for complete translation arrest are underlined. (B) CU164 (secY39) transformed with a plasmid encoding SecM(Eco), a SecM(Eco) mutant, or Sup1–6 were subjected to pulse-chase labeling after the addition of IPTG to induce expression of the plasmid-borne gene. Full-length and translation-arrested forms of SecM were immunoprecipitated and resolved by SDS-PAGE.

The plasticity of a translation arrest motif yields insights into nascent polypeptide recognition inside the ribosome tunnel