Wrapping of DNA around the E. coli RNA polymerase open promoter complex

Abstract

High-resolution atomic force microscopy (AFM) and biochemical methods were used to analyze the structure of Escherichia coli RNA polymerase·σ70 (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 ~ 30 nm (~ 90 bp) relative to the free DNA. The DNA bend angle measured with different methods varied from 55 to 88°. 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°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 σ70·Rp(O), conformation is proposed.