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Nucleic Acids Res. 2012 Aug;40(15):7442-51. doi: 10.1093/nar/gks383. Epub 2012 May 8.

Active site opening and closure control translocation of multisubunit RNA polymerase.

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1
Department of Biochemistry and Food Chemistry, University of Turku, 20014, Turku, Finland.

Abstract

Multisubunit RNA polymerase (RNAP) is the central information-processing enzyme in all cellular life forms, yet its mechanism of translocation along the DNA molecule remains conjectural. Here, we report direct monitoring of bacterial RNAP translocation following the addition of a single nucleotide. Time-resolved measurements demonstrated that translocation is delayed relative to nucleotide incorporation and occurs shortly after or concurrently with pyrophosphate release. An investigation of translocation equilibrium suggested that the strength of interactions between RNA 3' nucleotide and nucleophilic and substrate sites determines the translocation state of transcription elongation complexes, whereas active site opening and closure modulate the affinity of the substrate site, thereby favoring the post- and pre-translocated states, respectively. The RNAP translocation mechanism is exploited by the antibiotic tagetitoxin, which mimics pyrophosphate and induces backward translocation by closing the active site.

PMID:
22570421
PMCID:
PMC3424550
DOI:
10.1093/nar/gks383
[Indexed for MEDLINE]
Free PMC Article
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