NADH production from NAD+ using malic enzyme of Pseudomonas diminuta IFO-13182

Abstract Permeable cells and a sonicate of Pseudomonas diminuta IFO-13182 produced NADH from NAD − by an NAD- and NADP-linked malic enzyme [ l -malate: NAD(P) + oxidoreductase (EC 1.1.1.39)] reaction in the presence of l -malic acid and divalent metal ions. Optimum conditions for NADH production were examined, including malate concentration, NAD + concentration, pH, and buffer type. Under suitable conditions, the conversion ratio of NADH from NAD + reached 100% after 3 h of incubation at 30°C and the amount of NADH accumulated was 14 μmol ml −1 of reaction mixture. The NADH produced in such enzyme reaction was purified in powder form. The purity of NADH produced was 96.6% from its coenzyme activity with alcohol dehydrogenase.