Hydrogen-Dependent Nitrogenase Activity and ATP Formation in Rhizobium japonicum Bacteroids†

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Rhizobium japonicum 122 DES bacteroids from soybean nodules possess an active H2-oxidizing system that recycles all of the H2 lost through nitrogenase-dependent H2 evolution. The addition of 72 μM H2 to suspensions of bacteroids increased O2 uptake 300% and the rate of C2H2 reduction 300 to 500%. The optimal partial pressure of O2 was increased, and the partial pressure of O2 range for C2H2 reduction was extended by adding H2. A supply of succinate to bacteroids resulted in effects similar to those obtained by adding H2. Both H2 and succinate provided respiratory protection for the N2-fixing system in bacteroids. The oxidation of H2 by bacteroids increased the steady-state pool of ATP by 20 to 40%. In the presence of 50 mM iodoacetate, which caused much greater inhibition of endogenous respiration than of H2 oxidation, the addition of H2 increased the steady-state pool of ATP in bacteroids by 500%. Inhibitor evidence and an absolute requirement for O2 indicated that the H2-stimulated ATP synthesis occurred through oxidative phosphorylation. In the presence of 50 mM iodoacetate, H2-dependent ATP synthesis occurred at a rate sufficient to support nitrogenase activity. The addition of H2 to H2 uptake-negative strains of R. japonicum had no effect on ATP formation or C2H2 reduction. It is concluded that the H2-oxidizing system in H2 uptake-positive bacteroids benefits the N2-fixing process by providing respiratory protection of the O2-labile nitrogenase proteins and generating ATP to support maximal rates of C2H2 reduction by oxidation of the H2 produced from the nitrogenase system.

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