[Analysis of a new nitrogen fixation gene in Pseudomonas stutzeri A1501]

We studied the role of the nitrogen fixation gene PST1305 located within the nitrogen fixation island of Pseudomonas stutzeri A1501.

We constructed the mutant strain (np1305) by homologous recombination and triparental conjugation, and determined the nitrogenase activity by the acetylene reduction test. Through RT-PCR, we analyzed the transcriptional units of PST1305 gene and its nearby genes. Real-Time PCR was applied to compare the expression level of PST1305 gene between optimal and non-nitrogen fixating conditions.

Compared to the wild type, the nitrogenase activity in mutant strain (np1305) was partially decreased, however, functional complementary strain (np1305Comp) could restore nitrogenase activity close to wild type level. PST1305 gene was co-transcribed with its upstream gene (nifB and fdxN) and downstream gene (nifQ, PST1303 and PST1302). In contrast to the nitrogen excess conditions, expression of PST1305 under nitrogen-fixing conditions was significantly upregulated for 38.7-fold.

Disruption of PST1305 exhibited a declined nitrogenase activity compared to the wild type A1501. PST1305 gene might participate in biological nitrogen fixation by involving in the electron transport or the oxygen protection mechanism of nitrogenase. These results suggested that PST1305 gene was a new gene required for optimal nitrogenase activity of Pseudomonas stutzeri A1501.