In vivo and in vitro kinetics of nitrogenase

Publisher Website
Google Scholar
Abstract
We measured some of the kinetic parameters of nitrogenase in intact systems of Clostridium pasteurianum and Klebsiella pneumoniae to compared them with the kinetics of the enzyme in vitro. The enzyme showed mutiple apparent Km values for acetylene reduction in vivo, as it does in vitro. CO was a noncompetitive inhibitor of acetylene reduction; azide was a noncompetitive inhibitor of acetylene reduction, and nitrogen was a partial inhibitor of acetylene reduction. Cyanide was a noncompetitive inhibitor of acetylene reduction in C. pasteurianum but it was a metabolic poison in K. pneumoniae, in addition to being an inhibitor of nitrogenase. The partial nature of nitrogen inhibition was apparent in assays where both nitrogen and CO were present. Nitrogen did not alter the apparent Ki [inhibition constant] for CO, nor did the presence of CO enhance the competitive effectiveness of nitrogen. By using recombined nitrogenase fractions, we found that the ability of nitrogen to inhibit hydrogen evolution or acetylene reduction varied with the ratio of protein components. The in vivo inhibition of acetylene reduction by dinitrogen was comparable to that obtained with an excess of the Fe protein in vitro. There is apparently an effective excess of the Fe protein available under active growth conditions in vivo.