Isolation and sequence analysis of the GlnKamtB1amtB2 gene cluster, encoding a PII homologue and two putative ammonium transporters, from Pseudomonas stutzeri A15.

Abstract

By PCR, using primers based on heterologous amtB genes, an amtB sequence of Pseudomonas stutzeri A15 was amplified. This DNA fragment was used as a probe in Southern hybridisation experiments and resulted in the isolation and sequence analysis of a 6017 bp genomic fragment of P. stutzeri A15 containing glnKamtB1amtB2. GlnK codes for a homologue of the nitrogen regulatory PII protein, amtB1 and amtB2 encode putative ammonium transporters. Whereas a glnKamtB gene cluster is common among bacteria, a tandem repeat of ammonium transporter genes has not been reported before. Apart from the presence of a second amtB gene, the gene organisation on this 6 kbp fragment is very similar to a particular region in the genome of Pseudomonas aeruginosa PAO1, relatively closely related to P. stutzeri. Furthermore, the amtB1 gene shows the highest similarity with P. aeruginosa amtB, whereas the amtB2 gene is more closely related to cyanobacterial amtB genes, which are reported to be monocistronically transcribed and not clustered with glnK homologues. Upstream of glnK, NtrC and RpoN recognition sites can be observed. In the intergenic region of glnKamtB1amtB2 no terminators nor extra promoter sequences were observed, indicating that glnKamtB1amtB2 is possibly transcribed as a nitrogen regulated operon.