On nitroaryl reductase activities in several Clostridia.

Crude extracts of Clostridium kluyveri, Clostridium spec. La 1, Clostridium sporogenes and Clostridium pasteurianum catalyse the NADH-dependent reduction of the nitro group of p-nitrobenzoate. The former three Clostridia also use pyruvate as electron donor for this reduction. The NADH-dependent reductases have been partially purified and characterized from Clostridium kluyveri. Nitroalkyl compounds as well as nitrite, sulfite, sulfate and hydroxylamine are no substrates. Based on chromatographic behavior, separation pattern, yields, stability, pH optima, molecular masses and EPR studies the three NADH-dependent nitroaryl group reducing enzymes in Clostridium kluyveri (three activities in Clostridium spec. La 1 and two activities in Clostridium sporogenes) are different from alcohol dehydrogenase, aldehyde dehydrogenase, 3-hydroxy-butyryl-CoA dehydrogenase, butyryrl-CoA dehydrogenase, 2-enoate reductase, ferredoxin-NAD and ferredoxin-NADP reductase. The physiological roles of the nitroaryl reductases are not known. The reductase activities show losses of 80-90% during classical protein purification procedures. One of the three nitroaryl reductases exhibits a pH optimum of 10.5. The crude extract reveals a pH optimum at 11.5. The first step of the reduction reaction leads to the nitroradical anion (1 electron transfer). The electron transfer to p-nitrobenzoate is also catalysed by ferrodoxin-NAD reductase from NADH and by ferredoxin-NADP reductase from NADP. Partially purified 2-oxo-acid synthases from Clostridium sporogenes catalyse with low rates the reduction of p-nitrobenzoate as well as 2-nitroethanol in the presence and absence of ferredoxin using pyruvate or 2-oxo-4-methylpentanoate as electron donors, respectively. The NADH-dependent reduction of p-nitro-benzoate accounts for at least 70% and the 2-oxo acid-dependent reduction for about 5% of the total nitroaryl reductase activity in the Clostridia. It seems that the pyridine nucleotide-dependent nitroaryl reductases are enzymes so far unknown in Clostridia.