title: RNA polymerase stalls in a post-translocated register and can hyper-translocate.
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.
authors.affiliation: Department of Biochemistry and Molecular Biology, Michigan State University, East Lansing, MI, USA.
authors.forename: Yuri A
authors.initials: YA
authors.lastname: Nedialkov
authors.forename: Evgeny
authors.initials: E
authors.lastname: Nudler
authors.forename: Zachary F
authors.initials: ZF
authors.lastname: Burton
mesh_terms: D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D005090:Exodeoxyribonucleases / Q000378:metabolism*; D035602:Transcriptional Elongation Factors / Q000235:genetics / Q000378:metabolism; D014178:Translocation, Genetic
chemical_list: D029968:Escherichia coli Proteins; D035602:Transcriptional Elongation Factors; D012321:DNA-Directed RNA Polymerases; D005090:Exodeoxyribonucleases; C020133:exodeoxyribonuclease III
keywords: NTP analogues; RNA polymerase; bridge helix; hyper-translocation; tagetitoxin; translocation; trigger loop