Transcriptional activation of the Klebsiella pneumoniae nitrogenase promoter may involve DNA loop formation.

Transcriptional activation of nitrogen fixation genes by NifA in Klebsiella pneumoniae requires an upstream NifA binding site. We now report that the introduction of half turns of the DNA helix into the DNA separating the upstream NifA binding site from the downstream promoter element of the nifH promoter decreases NifA-mediated activation to a greater extent than does the introduction of full helical turns. Reducing the spacing between the upstream and downstream elements of the nifH promoter also results in a promoter down phenotype. Introduction of a tight protein-binding site, the lac operator, between the upstream and downstream promoter elements did not render activation of the nifH promoter sensitive to occupancy of this site by the lac repressor. These findings indicate that NifA-mediated activation of transcription requires that NifA is bound upstream, and to the correct face of the DNA helix, in order to interact with downstream transcription factors. This implies that the interaction is brought about by the formation of a DNA loop between upstream and downstream promoter elements rather than by NifA sliding downstream.