Comparision of Soluble Reduced Nicotinamide Adenine Dinucleotide Oxidases from Cells and Spores of Clostridium botulinum

Abstract

Green , J. H. (Michigan State University, East Lansing), and H. L. Sadoff . Comparison of soluble reduced nicotinamide adenine dinucleotide oxidases from cells and spores of Clostridium botulinum . J. Bacteriol. 89: 1499–1505. 1965.—The properties of purified reduced nicotinamide adenine dinucleotide (NADH 2 ) oxidases from cells and spores of Clostridium botulinum 62-A have been studied to determine whether they are the same or different proteins. The spore NADH 2 oxidase was very heat-stable, whereas the vegetative enzyme was readily denatured at 70 C. The spore oxidase exhibited less affinity for the substrate than did the vegetative protein, but possessed a tightly bound cofactor. Atabrine was a noncompetitive inhibitor for both enzymes, but was less inhibitory to the spore NADH 2 oxidase. The enzymes could be separated from each other by gel filtration or chromatography on a diethylaminoethyl-cellulose column. The molecular weight of the spore oxidase was estimated to be 200,000 or greater, whereas that of the vegetative enzyme was 100,000 or less. Neither NADH 2 oxidase would cross-react with its heterologous antibody in a precipitation reaction. The conclusion drawn from this investigation is that the two NADH 2 oxidases are distinctly different proteins.