Wrapping of DNA around the E.coli RNA polymerase open promoter complex.
High-resolution atomic force microscopy (AFM) and biochemical methods were used to analyze the structure of Escherichia coli RNA polymerase.sigma(70) (RNAP) open promoter complex (RP(o)). A detailed analysis of a large number of molecules shows that the DNA contour length of RP(o) is reduced by approximately 30 nm (approximately 90 bp) relative to the free DNA. The DNA bend angle measured with different methods varied from 55 to 88 degrees. The contour length reduction and the DNA bend angle were much less in inactive RNAP-DNA complexes. These results, together with previously published observations, strongly support the notion that during transcription initiation, the promoter DNA wraps nearly 300 degrees around the polymerase. This amount of DNA bending requires an energy of 60 kJ/mol. The structural analysis of the open promoter complexes revealed that two-thirds of the DNA wrapped around the RNAP is part of a region upstream of the transcription start site, whereas the remaining one-third is part of the downstream region. Based on these data, a model of the sigma(70).RP(o) conformation is proposed.
Rivetti
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Istituto di Scienze Biochimiche, Università di Parma, 43100 Parma, Italy.
Guthold
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Bustamante
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D001426:Bacterial Proteins / Q000378:metabolism*; D010582:Bacteriophage lambda / Q000235:genetics*; D004279:DNA, Viral / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D018625:Microscopy, Atomic Force; D009532:Nickel; D009690:Nucleic Acid Conformation; D011401:Promoter Regions, Genetic; D012808:Sigma Factor / Q000378:metabolism*; D013698:Templates, Genetic
D001426:Bacterial Proteins; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D012808:Sigma Factor; C029938:nickel sulfate; D009532:Nickel; C081775:RNA polymerase sigma 70; D012321:DNA-Directed RNA Polymerases