title: Thio-Modification of tRNA at the Wobble Position as Regulator of the Kinetics of Decoding and Translocation on the Ribosome.
abstract: Uridine 34 (U34) at the wobble position of the tRNA anticodon is post-transcriptionally modified, usually to mcm5s2, mcm5, or mnm5. The lack of the mcm5or s2modification at U34of tRNALys, tRNAGlu, and tRNAGlncauses ribosome pausing at the respective codons in yeast. The pauses occur during the elongation step, but the mechanism that triggers ribosome pausing is not known. Here, we show how the s2modification in yeast tRNALysaffects mRNA decoding and tRNA-mRNA translocation. Using real-time kinetic analysis we show that mcm5-modified tRNALyslacking the s2group has a lower affinity of binding to the cognate codon and is more efficiently rejected than the fully modified tRNALys. The lack of the s2modification also slows down the rearrangements in the ribosome-EF-Tu-GDP-Pi-Lys-tRNALyscomplex following GTP hydrolysis by EF-Tu. Finally, tRNA-mRNA translocation is slower with the s2-deficient tRNALys. These observations explain the observed ribosome pausing at AAA codons during translation and demonstrate how the s2modification helps to ensure the optimal translation rates that maintain proteome homeostasis of the cell.
authors.affiliation: Department of Physical Biochemistry, Max Planck Institute for Biophysical Chemistry , Am Fassberg 11, 37077 Goettingen, Germany.
authors.forename: Namit
authors.initials: N
authors.lastname: Ranjan
authors.affiliation: Department of Physical Biochemistry, Max Planck Institute for Biophysical Chemistry , Am Fassberg 11, 37077 Goettingen, Germany.
authors.forename: Marina V
authors.identifier: 0000-0003-0105-3879
authors.initials: MV
authors.lastname: Rodnina
mesh_terms: D007700:Kinetics; D012343:RNA, Transfer / Q000737:chemistry / Q000378:metabolism*; D012270:Ribosomes / Q000737:chemistry / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000737:chemistry / Q000378:metabolism*; D014529:Uridine / Q000737:chemistry / Q000378:metabolism
chemical_list: D013438:Sulfhydryl Compounds; D012343:RNA, Transfer; D014529:Uridine