RNA polymerase stalls in a post-translocated register and can hyper-translocate

Transcription. Sep-Oct 2012;3(5):260-9. doi: 10.4161/trns.22307. Epub 2012 Sep 1.

Abstract

Exonuclease (Exo) III was used to probe translocation states of RNA polymerase (RNAP) ternary elongation complexes (TECs). Escherichia coli RNAP stalls primarily in a post-translocation register that makes relatively slow excursions to a hyper-translocated state or to a pre-translocated state. Tagetitoxin (TGT) strongly inhibits hyper-translocation and inhibits backtracking, so, as indicated by Exo III mapping, TGT appears to stabilize both the pre- and probably a partially post-translocation state of RNAP. Because the pre-translocated to post-translocated transition is slow at many template positions, these studies appear inconsistent with a model in which RNAP makes frequent and rapid (i.e., millisecond phase) oscillations between pre- and post-translocation states. Nine nucleotides (9-nt) and 10-nt TECs, and TECs with longer nascent RNAs, have distinct translocation properties consistent with a 9-10 nt RNA/DNA hybrid. RNAP mutant proteins in the bridge helix and trigger loop are identified that inhibit or stimulate forward and backward translocation.

Keywords: NTP analogues; RNA polymerase; bridge helix; hyper-translocation; tagetitoxin; translocation; trigger loop.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • DNA-Directed RNA Polymerases / genetics
  • DNA-Directed RNA Polymerases / metabolism*
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Escherichia coli Proteins / genetics
  • Escherichia coli Proteins / metabolism*
  • Exodeoxyribonucleases / metabolism*
  • Transcriptional Elongation Factors / genetics
  • Transcriptional Elongation Factors / metabolism
  • Translocation, Genetic

Substances

  • Escherichia coli Proteins
  • Transcriptional Elongation Factors
  • DNA-Directed RNA Polymerases
  • Exodeoxyribonucleases
  • exodeoxyribonuclease III