Nitrogenase activity in Rhodopseudomonas sulfidophila

The marine purple nonsulfur bacterium, Rhodopseudomonas sulfidophila, strain W4, was capable of photosynthetic growth on dinitrogen and malate. Higher growth rates were observed when either glutamate or ammonia replaced dinitrogen as nitrogen source and when bicarbonate was omitted from the culture medium. Although ammonia was released from cells growing on malate and N₂, no nitrogenase activity could be detected unless α-ketoglutarate was added to the culture medium. No nitrogenase activity was found in cultures grown in the presence of NH₄⁺. In cultures grown on glutamate as nitrogen source, nitrogenase and hydrogenase activities were found to be 5.4 nmol C₂H₂ reduced · min^-1 · mg^-1 dry weight and 50 nmol methylene blue reduced · min^-1 · mg^-1 dry weight respectively. Such activities are significantly lower than those observed for other members of the Rhodospirillaceae e.g. Rhodopseudomonas capsulata. However, the hydrogenase activity would be sufficient to recycle all H₂ produced by nitrogenase. It was indeed observed that growing cells did not evolve molecular hydrogen during photoheterotrophic growth and that H₂ stimulated nitrogenase activity in resting cells of R. sulfidophila. The nitrogenase from this bacterium proved to be extremely sensitive to low concentrations of oxygen, half-inhibition occurring at between 1–1.5% O₂ in the gas phase, depending on the bacterial concentration. Light was essential for nitrogenase activity. No activity was found during growth in the dark under extremely low oxygen concentrations (1–2% O₂), which are still sufficient to support good growth. Resting cell suspensions prepared from such cultures were unable to reduce acetylene upon illumination. Optimum nitrogenase activities were broadly defined over the temperature range, 30–38°C, and between pH 6.9 and 8.0. The results are discussed in comparison with the non-marine purple nonsulfur bacterium, R. capsulata, which somewhat resembles R. sulfidophila.