A novel dye-linked alcohol dehydrogenase activity present in some Gram-positive bacteria

An assay was developed which allowed reproducible detection of methanol oxidation by cell free extracts of methanol-grown Amycolatopsis methanolica. The dye-linked activity was only observed when high concentrations of phosphate or sulphate salts were applied in the assay. The specific activity strongly increased when raising the amount of cell free extract in the assay. From the large number of electron acceptors tested only the tetrazolium dyes MTT and INT showed significant activity. Activity was observed with primary as well as secondary alcohols. A similar activity was found in ethanol-grown A. methanolica, Rhodococcus erythropolis and Rhodococcus rhodochrous, but not in other bacteria tested. The identity and possible involvement in alcohol oxidation of this novel dye-linked activity is discussed.

Formaldehyde dismutase activities in Gram-positive bacteria oxidizing methanol

Extracts of methanol-grown cells of Amycolatopsis methanolica and Mycobacterium gastri oxidized methanol and ethanol with concomitant reduction of N,N’-dimethyl-4-nitrosoaniline (NDMA). Anion-exchange chromatography revealed the presence of a single enzyme able to catalyse this activity in methanol- or ethanol-grown cells of M. gastri. A. methanolica, however, possessed two different enzymes, one of which was similar to the single enzyme found in M. gastri. The methanol:NDMA oxidoreductases (MNO) were purified to homogeneity from methanol-grown cells of A. methanolica and M. gastri. Both enzyme preparations showed similar relative molecular masses with subunits of Mr 50000 and 49000, and native enzymes of Mr 268000 and 255000 (gel-filtration data for A. methanolica and M. gastri, respectively). Both enzymes also displayed a similar substrate specificity. They were active with methanol and various other primary alcohols (yielding the corresponding aldehydes), polyols and formaldehyde. In addition, the MNO enzymes produced methylformate from methanol plus formaldehyde, and catalyzed formaldehyde dismutase and NADH-dependent formaldehyde reductase reactions. They did not possess NAD(P)+- or dye-linked alcohol dehydrogenase or oxidase activities.