title: Peptide release promoted by methylated RF2 and ArfA in nonstop translation is achieved by an induced-fit mechanism.
abstract: Here we report that the specificity of peptide release in the ribosome on a nonstop mRNA by ArfA and RF2 is achieved by an induced-fit mechanism. Using RF2 that is methylated on the glutamine of its GGQ motif (RF2(m)), we show that methylation substantially increases the rate of ArfA/RF2-catalyzed peptide release on a nonstop mRNA that does not occupy the ribosomal A site, but has only a modest effect on k(cat) by the same proteins on longer nonstop mRNAs occupying the A site of the mRNA channel in the ribosome. Our data suggest that enhancement in the kcat of peptide release by ArfA and RF2 under the cognate decoding condition is the result of favorable conformational changes in the nonstop complex. We demonstrate a shared mechanism between canonical and nonstop termination, supported by similarities in the kinetic mechanisms in antibiotic inhibition and methylation-correlated enhancement in the rate of peptide release. Despite these similarities, our data suggest that nonstop termination differs from canonical pathway in the downstream event of recycling.
authors.affiliation: Department of Biochemistry, Center for Biophysics and Quantitative Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
authors.forename: Fuxing
authors.initials: F
authors.lastname: Zeng
authors.affiliation: Department of Biochemistry, Center for Biophysics and Quantitative Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA hjin@illinois.edu.
authors.forename: Hong
authors.initials: H
authors.lastname: Jin
mesh_terms: D055162:Biocatalysis; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000378:metabolism*; D008745:Methylation; D010303:Paromomycin / Q000494:pharmacology; D010443:Peptide Chain Termination, Translational; D010454:Peptide Termination Factors / Q000378:metabolism*; D010455:Peptides / Q000378:metabolism*; D014176:Protein Biosynthesis; D016601:RNA-Binding Proteins / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism; D014756:Viomycin / Q000494:pharmacology
chemical_list: C556014:ArfA protein, E coli; D029968:Escherichia coli Proteins; D010454:Peptide Termination Factors; D010455:Peptides; D016601:RNA-Binding Proteins; C489006:prfB protein, E coli; D010303:Paromomycin; D014756:Viomycin
keywords: RF2; alternative rescuing factor A; nonstop translation; quality control; ribosome; translational termination