Effects of Disruption of Homocitrate Synthase Genes on Nostoc sp. Strain PCC 7120 Photobiological Hydrogen Production and Nitrogenase

Abstract

ABSTRACT In the case of nitrogenase-based photobiological hydrogen production systems of cyanobacteria, the inactivation of uptake hydrogenase (Hup) leads to significant increases in hydrogen production activity. However, the high-level-activity stage of the Hup mutants lasts only a few tens of hours under air, a circumstance which seems to be caused by sufficient amounts of combined nitrogen supplied by active nitrogenase. The catalytic FeMo cofactor of nitrogenase binds homocitrate, which is required for efficient nitrogen fixation. It was reported previously that the nitrogenase from the homocitrate synthase gene ( nifV ) disruption mutant of Klebsiella pneumoniae shows decreased nitrogen fixation activity and increased hydrogen production activity under N 2 . The cyanobacterium Nostoc sp. strain PCC 7120 has two homocitrate synthase genes, nifV1 and nifV2 , and with the Δ hupL variant of Nostoc sp. strain PCC 7120 as the parental strain, we have constructed two single mutants, the Δ hupL Δ nifV1 strain (with the hupL and nifV1 genes disrupted) and the Δ hupL Δ nifV2 strain, and a double mutant, the Δ hupL Δ nifV1 Δ nifV2 strain. Diazotrophic growth rates of the two nifV single mutants and the double mutant were decreased moderately and severely, respectively, compared with the rates of the parent Δ hupL strain. The hydrogen production activity of the Δ hupL Δ nifV1 mutant was sustained at higher levels than the activity of the parent Δ hupL strain after about 2 days of combined-nitrogen step down, and the activity in the culture of the former became higher than that in the culture of the latter. The presence of N 2 gas inhibited hydrogen production in the Δ hupL Δ nifV1 Δ nifV2 mutant less strongly than in the parent Δ hupL strain and the Δ hupL Δ nifV1 and Δ hupL Δ nifV2 mutants. The alteration of homocitrate synthase activity can be a useful strategy for improving sustained photobiological hydrogen production in cyanobacteria.