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PMID: 16284617 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, U.S. Gov't, Non-P.H.S.

Direct observation of base-pair stepping by RNA polymerase.

Nature ·Vol. 438 ·No. 7067 ·2005-11-24 ·Pages 460-5

Abbondanzieri EA, Greenleaf WJ, Shaevitz JW, Landick R, Block SM

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.

MeSH Terms
Base Pairing DNA/chemistry,genetics,metabolism DNA-Directed RNA Polymerases/metabolism Escherichia coli/enzymology Kinetics Models, Biological Movement Nucleotides/genetics,metabolism Optics and Photonics Sensitivity and Specificity Templates, Genetic Transcription, Genetic
Chemicals
Nucleotides DNA DNA-Directed RNA Polymerases
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Abbondanzieri Elio A
Department of Applied Physics, Stanford University, Stanford, California 94305, USA.
Greenleaf William J
Shaevitz Joshua W
Landick Robert
Block Steven M
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Article Info
Journal
Nature
Abbr.
Nature
ISSN
1476-4687
DOI
10.1038/nature04268
Published
2005-11-24
Epub
2005-00-13
Pages
460-5
Language
English
Region
England
NLM ID
0410462
PMCID
PMC1356566
Subset
IM
Grants
NIGMS NIH HHS · R01 GM057035 · United States
External Links
PubMed source DOI: 10.1038/nature04268 PMC Full Text View journal details
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