COMPARISON AND CROSS REACTIONS OF NITROGENASE FROM KLEBSIELLA-PNEUMONIAE, AZOTOBACTER-CHROOCOCCUM AND BACILLUS-POLYMYXA

Crude nitrogenase of Klebsiella pneumoniae was less rapidly inactivated by 0.2 atm of O2 than nitrogenase of Bacillus polymyxa; inactivation of Azotobacter chroococcum nitrogenase required 1 atm of O2. Nitrogenase from each organism was separated into two protein components; one was rapidly inactivated by air, the other was only slowly affected. The product of reduction of C2H2 in 2H2O or C22H2 in H2O by each nitrogenase was cis-ethylene, C22H2H2, some trans-C22H2H2 was also detected. Each nitrogenase catalysed exchange between 2H2 and H2O proportional to the partial pressure of 2H2 and not dependent on the presence of N2. Proteins 1 of each nitrogenase were assayed with their corresponding Protein 2 or those of the other two bacteria for rate of acetylene, azide, cyanide, isocyanide or N2 reduction; ATP-dependent H2 evolution and Pi formed from ATP hydrolysis were also determined. Cross reaction, better than 80%, was observed between Azotobacter chroococcum and Klebsiella pneumoniae and between Bacillus polymyxa and Klebsiella pneumoniae with acetylene as substrate, but in the crosses Bacillus polymyxa Protein Protein 1 + Azotobacter chroococcum Protein 2 and its reciprocal only 50 and 12% cross, respectively, were observed. Some differences were found in the degree of cross reaction with the various substrates and the amount of Pi formed did not always correspond with the amount of substrate reduced. These results together with other work are used to support the hypothesis that cyanide or isocyanide are not model substrates, being reduced at more than one site on the nitrogenase complex. The possibility that a two-metal site catalyses N2 fixation is considered. © 1969.