title: EF-G catalyzed translocation dynamics in the presence of ribosomal frameshifting stimulatory signals.
abstract: Programmed -1 ribosomal frameshifting (-1PRF) is tightly regulated by messenger RNA (mRNA) sequences and structures in expressing two or more proteins with precise ratios from a single mRNA. Using single-molecule fluorescence resonance energy transfer (smFRET) between (Cy5)EF-G and (Cy3)tRNALys, we studied the translational elongation dynamics of -1PRF in the Escherichia coli dnaX gene, which contains three frameshifting signals: a slippery sequence (A AAA AAG), a Shine-Dalgarno (SD) sequence and a downstream hairpin. The frameshift promoting signals mostly impair the EF-G-catalyzed translocation step of the two tRNALys and the slippery codons from the A- and P- sites. The hairpin acts as a road block slowing the translocation rate. The upstream SD sequence together with the hairpin promotes dissociation of futile EF-G and thus causes multiple EF-G driven translocation attempts. A slippery sequence also helps dissociation of the EF-G by providing alternative base-pairing options. These results indicate that frameshifting takes place during the repetitive ribosomal conformational changes associated with EF-G dissociation upon unsuccessful translocation attempts of the second slippage codon from the A- to the P- sites.
authors: Department of Chemistry, University of California, Berkeley, CA 94720, USA.
authors: Hee-Kyung
authors: HK
authors: Kim
authors: Department of Chemistry, University of California, Berkeley, CA 94720, USA.
authors: Ignacio
authors: I
authors: Tinoco
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics; D055162:Biocatalysis; D003062:Codon; D004258:DNA Polymerase III / Q000235:genetics; D031541:Fluorescence Resonance Energy Transfer; D018965:Frameshifting, Ribosomal; D009154:Mutation; D010441:Peptide Chain Elongation, Translational; D020653:Peptide Elongation Factor G / Q000378:metabolism*; D012333:RNA, Messenger / Q000737:chemistry; D012357:RNA, Transfer, Lys / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D003062:Codon; C103076:DnaX protein, Bacteria; D020653:Peptide Elongation Factor G; D012333:RNA, Messenger; D012357:RNA, Transfer, Lys; D004258:DNA Polymerase III