The Metabolism of Nicotinic Acid

Purification and crystallization of an enzyme which catalyzes the oxidation of 2,5-dihydroxypyridine, an intermediate in nicotinic acid catabolism, are described. The labile enzyme is stabilized by dithiothreitol. Activity lost by dialysis or purification procedures is restored by incubation with dithiothreitol and ferrous sulfate. The enzyme is inhibited by sulfhydryl reagents and iron-chelating agents. The crystalline enzyme, on polyacrylamide electrophoresis with dithiothreitol, yields a single major band and a region of minor diffuse bands. In the absence of dithiothreitol the intensity of the minor bands is increased. On acrylamide gels containing sodium dodecyl sulfate, a single band with mobility corresponding to a molecular weight of 39,500 is obtained. A single region of enzyme activity corresponding to a molecular weight of 242,000 is obtained on sucrose gradients containing dithiothreitol. A portion of this activity is shifted to a region corresponding to lower molecular weight when dithiothreitol is omitted.