Photochemical cross-linking studies on the interaction of Escherichia coli RNA polymerase with T7 DNA

Abstract

We have identified the subunits of Escherichia coli RNA polymerase which are in close contact with the T7 phage DNA template using photochemical cross-linking. In nonspecific T7 DNA-enzyme complexes which occur in all regions of the DNA, subunits sigma, beta, and beta' were cross-linked to the DNA. In contrast, in specific binary complexes which presumably occur at promoter sites, and in the initiation complex (holoenzyme + T7 DNA + initiator dinucleotides + three nucleoside triphosphates), only sigma and beta were cross-linked to DNA, while cross-linking of beta' could not be demonstrated. These results (1) do not support the idea that alpha subunits are involved in the enzyme-template interaction, (2) raise the possibility that sigma subunit participates directly in promoter recognition even though isolated sigma does not bind to DNA, and (3) indicate different modes of interaction between RNA polymerase and DNA in nonspecific and specific complexes. These findings are relevant to the mechanism by which RNA polymerase carries out selective transcription.