A new tRNA intermediate revealed on the ribosome during EF4-mediated back-translocation

Nat Struct Mol Biol. 2008 Sep;15(9):910-5. doi: 10.1038/nsmb.1469.

Abstract

EF4 (LepA) is an almost universally conserved translational GTPase in eubacteria. It seems to be essential under environmental stress conditions and has previously been shown to back-translocate the tRNAs on the ribosome, thereby reverting the canonical translocation reaction. In the current work, EF4 was directly visualized in the process of back-translocating tRNAs by single-particle cryo-EM. Using flexible fitting methods, we built a model of ribosome-bound EF4 based on the cryo-EM map and a recently published unbound EF4 X-ray structure. The cryo-EM map establishes EF4 as a noncanonical elongation factor that interacts not only with the elongating ribosome, but also with the back-translocated tRNA in the A-site region, which is present in a previously unseen, intermediate state and deviates markedly from the position of a canonical A-tRNA. Our results, therefore, provide insight into the underlying structural principles governing back-translocation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biological Transport, Active
  • Escherichia coli / metabolism
  • Escherichia coli Proteins / chemistry*
  • Escherichia coli Proteins / metabolism*
  • Guanosine Triphosphate / metabolism
  • Hydrolysis
  • Macromolecular Substances
  • Models, Molecular
  • Peptide Initiation Factors
  • RNA, Bacterial / chemistry*
  • RNA, Bacterial / metabolism*
  • RNA, Transfer / chemistry*
  • RNA, Transfer / metabolism*
  • Ribosomes / chemistry*
  • Ribosomes / metabolism*
  • Transcriptional Elongation Factors / chemistry*
  • Transcriptional Elongation Factors / metabolism*

Substances

  • Escherichia coli Proteins
  • LepA protein, E coli
  • Macromolecular Substances
  • Peptide Initiation Factors
  • RNA, Bacterial
  • Transcriptional Elongation Factors
  • Guanosine Triphosphate
  • RNA, Transfer

Associated data

  • PDB/1524