CATABOLISM OF ISONICOTINATE BY MYCOBACTERIUM SP INA1 - EXTENDED DESCRIPTION OF THE PATHWAY AND PURIFICATION OF THE MOLYBDOENZYME ISONICOTINATE DEHYDROGENASE

Catabolism of isonicotinate by Mycobacterium sp. INA1 has been shown t o proceed via 2-hydroxyisonicotinate, 2,6-dihydroxyisonicotinate (citr azinate), citrazyl-CoA and 2,6-dioxopiperidine-4-carboxyl-CoA. An exte nded pathway involving propane-1,2,3-tricarboxylate as a further inter mediate is presented in this paper. Propane-1,2,3-tricarboxylate was o xidized stepwise to 2-oxoglutarate involving an oxidase, aconitase and isocitrate dehydrogenase. Isonicotinate dehydrogenase catalyses the f irst step of isonicotinate metabolism in Mycobacterium sp. INA1. The e nzyme was purified to apparent homogeneity by a three-step procedure. Enrichment was accompanied by partial loss in specific activity. The n ative enzyme had a molecular mass of either 125 kDa or 250 kDa, when e stimated by native gradient PAGE or gel filtration, respectively. SDS- gel electrophoresis revealed three types of subunits with molecular ma sses of approximately 83, 31 and 19 kDa. N-Terminal amino acid sequenc es of all three subunits have been determined. Molybdenum, iron, acid- labile sulphur and FAD were present at molar ratios of 1, 4, 4, 1 per protomer (125 kDa). The molybdenum-complexing cofactor was shown to be molybdopterin cytosine dinucleotide. Besides isonicotinate, only quin oline-4-carboxylate was found to be oxidized at appreciable rates.