Ribose 2′-hydroxyl groups in the 5′ strand of the acceptor arm of P-site tRNA are not essential for EF-G catalyzed translocation

Translocation of fragmented tRNA^Met from the ribosomal P-site. (A) Translocation of fragmented tRNA^Met having either unsubstituted or fully 2′-O-methyl–substituted small fragment. (Lane (−) RNA 1–17) Absence of the 5′ small fragment, (lane RNA 1–17) presence of unsubstituted 5′ small fragment, (lane 2′-O-me 1–17) presence of fully 2′-O-methyl-substituted small fragment. Minus (−) and plus (+) signs indicate absence or presence of EF-G. Arrows indicate toeprints before (Pre) and after (Post) translocation. (B) Translocation of fragmented tRNA^Met having single 2′-deoxy substitutions at positions 1–17 in the small fragment. Lanes as indicated above. (Lane tRNA_f^Met) Native E. coli tRNA_f^Met. (C) Extent of translocation of fragmented tRNA^Met with single 2′-deoxy substitution at the indicated positions. Values were normalized with respect to translocation of the unsubstituted fragmented tRNA^Met, which was set to 100%. Results are the average of three experiments.