nitrogenase and formic-acid

The reduction of N2 to NH3 by Mo-dependent nitrogenase at its active-site metal cluster FeMo-cofactor utilizes reductive elimination of Fe-bound hydrides with obligatory loss of H2 to activate the enzyme for binding/reduction of N2. Earlier work showed that wild-type nitrogenase and a nitrogenase with amino acid substitutions in the MoFe protein near FeMo-cofactor can catalytically reduce CO2 by two or eight electrons/protons to carbon monoxide (CO) and methane (CH4) at low rates. Here, it is demonstrated that nitrogenase preferentially reduces CO2 by two electrons/protons to formate (HCOO(-)) at rates >10 times higher than rates of CO2 reduction to CO and CH4. Quantum mechanical calculations on the doubly reduced FeMo-cofactor with a Fe-bound hydride and S-bound proton (E2(2H) state) favor a direct reaction of CO2 with the hydride ("direct hydride transfer" reaction pathway), with facile hydride transfer to CO2 yielding formate. In contrast, a significant barrier is observed for reaction of Fe-bound CO2 with the hydride ("associative" reaction pathway), which leads to CO and CH4. Remarkably, in the direct hydride transfer pathway, the Fe-H behaves as a hydridic hydrogen, whereas in the associative pathway it acts as a protic hydrogen. MoFe proteins with amino acid substitutions near FeMo-cofactor (α-70(Val→Ala), α-195(His→Gln)) are found to significantly alter the distribution of products between formate and CO/CH4.

Topics: Azotobacter vinelandii; Carbon Dioxide; Carbon Monoxide; Formates; Methane; Molybdoferredoxin; Nitrogenase; Oxidation-Reduction

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.

Topics: Bacillus; Carbon Dioxide; Clostridium; Formates; Hydrogen; Hydrogenase; Nitrogenase; Oxidation-Reduction; Oxidoreductases; Paenibacillus; Plasmodiophorida; Pyruvates