Nitrogenase. VII. Effect of component ratio, ATP and H2 on the distribution of electrons to alternative substrates.
Some kinetic properties of purified component I (Mo-Fe protein) and component II (Fe protein) of nitrogenase (EC 1.7.99.2) from Azotobacter vinelandii have been examined. The apparent Km values for reducible substrates (0.1 atm for N2, 0.01 atm for acetylene) and dithionite (0.5 mM) are similar for osmotically shocked cell lysates and purified components. However, the ATP dependence of acetylene and N2 reduction varies sigmoidally with ATP concentration and as a function of the relative and absolute concentration of components I and II in the assay. Acetylene is reduced in preference to N2 in competitive assays when component I is in relative excess. Acetylene reduction is not as dependent upon ATP concentration as is N2 reduction, so that acetylene is also a preferred substrate at lower ATP levels. Hydrogen specifically inhibits N2 reduction, diverting electrons to acetylene when both substrates are present in the assay. We propose a model of the enzyme activity, in which the substrates for reduction are bound to component I with electrons being activated by component II. ATP may be involved in activating electrons and in maintaining the appropriate conformation or reduction state of components to allow effective reduction of substrates. The relative rate of reduction of alternative substrates is dependent on the concentration of the particular state(s) capable of reacting with each substrate. The concentration of a particular state of component I is a function of components I, II and ATPL
D000114:Acetylene / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology*; D001395:Azotobacter / Q000201:enzymology; D004227:Dithionite / Q000494:pharmacology; D004583:Electrons; D006859:Hydrogen / Q000494:pharmacology*; D007700:Kinetics; D009591:Nitrogenase / Q000378:metabolism*; D010084:Oxidation-Reduction; D011485:Protein Binding
D004227:Dithionite; D006859:Hydrogen; D000255:Adenosine Triphosphate; D009591:Nitrogenase; D000114:Acetylene