The role of activator binding sites in transcriptional control of the divergently transcribed nifF and nifLA promoters from Klebsiella pneumoniae

Abstract

The regulatory region spanning the divergently transcribed nifF and nifLA promoters contains a NIFA-specific upstream activator sequence (UAS) located around +59, and two NTRC binding sites centred at -142 and -163 with respect to the nifLA transcription start site. We have constructed mutations in each of these binding sites and examined their role in transcriptional activation of the divergently transcribed promoters. Analysis of a mutation at +60 confirms that the UAS is required for efficient NIFA-mediated activation of nifF transcription. This sequence is also required for maximal activation of the nifLA promoter. Mutations at -169 and -148, within the two NTRC binding sites, reduce activation of the nifLA promoter by NTRC in vivo and lower the affinity of the activator for these sites in vitro. Phosphorylation of NTRC by NTRB is required for efficient binding of NTRC to these sites.