Direct observation of base-pair stepping by RNA polymerase. Author Elio Abbondanzieri, William Greenleaf, Joshua Shaevitz, Robert Landick, Steven Block Publication Year 2005 Type Journal Article 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. Keywords Escherichia coli, Transcription, Genetic, Models, Biological, DNA, Movement, Kinetics, Sensitivity and Specificity, Nucleotides, DNA-Directed RNA Polymerases, Base Pairing, Optics and Photonics, Templates, Genetic Journal Nature Volume 438 Issue 7067 Pages 460-5 Date Published 11/2005 Alternate Journal Nature Google ScholarBibTeXEndNote X3 XML