A mutant RNA polymerase that forms unusual open promoter complexes

We describe a mutant Escherichia coli RNA polymerase (RNAP) that forms stable open promoter complexes even at −20°C but with a shortened melted region that extends downstream to only position −7. In the presence of initiating transcription substrates, the mutant RNAP undergoes a temperature-dependent isomerization, resulting in a promoter complex that is indistinguishable from the wild-type RNAP–promoter complex, with the melted region extended downstream to position +4. We propose that the open complex formed by the mutant RNAP represents an intermediate on the normal promoter-opening pathway and that our results support earlier findings that initial promoter opening occurs in the upstream region of the −10 promoter consensus element and subsequently extends downstream to encompass the transcription start site.