Regulation of glutamate dehydrogenase expression in Pseudomonas putida results from its direct repression by NtrC under nitrogen-limiting conditions.

Nitrogen-regulated genes in enterobacteria are positively controlled by the transcriptional activator of σ(N) -dependent promoters NtrC, either directly or indirectly, through the dual regulator Nac. Similar to enterobacteria, gdhA encoding glutamate dehydrogenase from Pseudomonas putida is one of the few genes that is induced by excess nitrogen. In P. putida, the binding of NtrC to the gdhA promoter region and in vitro transcription suggest that, unlike its enterobacterial homologue that is repressed by Nac, gdhA is directly repressed by NtrC. Footprinting analyses demonstrated that NtrC binds to four distinct sites in the gdhA promoter. NtrC dimers bind cooperatively, and those bound closer to the promoter interact with the dimers bound further upstream, thus producing a proposed repressor loop in the DNA. The formation of the higher-order complex and the repressor loop appears to be important for repression but not absolutely essential. Both the phosphorylated and the non-phosphorylated forms of NtrC efficiently repressed gdhA transcription in vitro and in vivo. Therefore, NtrC repression of gdhA under nitrogen-limiting conditions does not depend on the phosphorylation of the regulator; rather, it relies on an increase in the repressor concentration under these conditions.