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Nature. 2005 Nov 24;438(7067):460-5. Epub 2005 Nov 13.

Direct observation of base-pair stepping by RNA polymerase.

Author information

1
Department of Applied Physics, Stanford University, Stanford, California 94305, USA.

Abstract

During transcription, RNA polymerase (RNAP) moves processively along a DNA template, creating a complementary RNA. Here we present the development of an ultra-stable optical trapping system with ångström-level resolution, which we used to monitor transcriptional elongation by single molecules of Escherichia coli RNAP. Records showed discrete steps averaging 3.7 +/- 0.6 A, a distance equivalent to the mean rise per base found in B-DNA. By combining our results with quantitative gel analysis, we conclude that RNAP advances along DNA by a single base pair per nucleotide addition to the nascent RNA. We also determined the force-velocity relationship for transcription at both saturating and sub-saturating nucleotide concentrations; fits to these data returned a characteristic distance parameter equivalent to one base pair. Global fits were inconsistent with a model for movement incorporating a power stroke tightly coupled to pyrophosphate release, but consistent with a brownian ratchet model incorporating a secondary NTP binding site.

PMID:
16284617
PMCID:
PMC1356566
DOI:
10.1038/nature04268
[Indexed for MEDLINE]
Free PMC Article

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