HYDROGENASE AND NITROGENASE IN CELL-FREE EXTRACTS OF BACILLUS POLYMYXA

Grau, F. H. (University of Wisconsin, Madison), and P. W. Wilson. Hydrogenase and nitrogenase in cell-free extracts of Bacillus polymyxa. J. Bacteriol. 85:446-450. 1963.-Washed cells of Bacillus polymyxa strain Hino, treated with lysozyme, yield cell-free extracts that rapidly evolve hydrogen from reduced methyl viologen, formate, and pyruvate. Hydrogenase is particulate, 86% being sedimented at 105,000 x g for 60 min. About 65% of the pyruvate metabolized is oxidized to acetyl phosphate, hydrogen, and carbon dioxide; the rest is converted to acetoin. These extracts fix considerable amounts of N(2) (15) when pyruvate is supplied as substrate, but will not fix with formate or mannitol. Centrifugation studies, and the absence of fixation with mannitol, show that this fixation is not caused by residual whole cells or spheroplasts. Cell-free fixation by B. polymyxa is similar to that by Clostridium pasteurianum. A short time lag in fixation occurs, and an optimal concentration of pyruvate is needed for maximal fixation. Arsenate causes a strong inhibition of fixation, presumably because arsenolysis of acetyl phosphate makes high-energy phosphate unavailable for the fixation process.