Nicotinoprotein [NAD(P)-containing] alcohol/aldehyde oxidoreductases. Purification and characterization of a novel type from Amycolatopsis methanolica.
Extracts of Gram-positive bacteria like Rhodococcus rhodochrous, Rhodococcus erythropolis and Amycolatopsis methanolica, but not those of several Gram-negative ones, showed dehydrogenase activity for ethanol as well as for methanol when 4-nitroso-N,N-dimethylaniline (NDMA) was used as electron acceptor. Chromatography of extracts of the first two organisms revealed one activity for both substrates, that of A. methanolica two activities, one of which is able to oxidize methanol and has been purified (Bystrykh, L.V., Govorukhina, N.I., van Ophem, P.W., Hektor, H.J., Dijkhuizen, L. and Duine, J.A., unpublished results). The other, indicated as NDMA-dependent alcohol dehydrogenase (NDMA-ADH), was purified to homogeneity. It is a trimeric enzyme consisting of subunits of 39 kDa and one firmly bound NAD as cofactor. Although NDMA-ADH shows structural similarity with the long-chain, zinc-containing, NAD(P)-dependent alcohol dehydrogenases with respect to the N-terminal sequence up to residue 41 (56% identity with horse liver alcohol dehydrogenase), the enzymes are catalytically different since NDMA-ADH is unable to use NAD(P)(H) as a coenzyme and NAD(P)-dependent alcohol dehydrogenases are inactive with NDMA (in the absence of NAD). Comparison of the NDMA-ADH properties with those of the methanol-oxidizing enzyme of A. methanolica, Mycobacterium gastri and Bacillus methanolica C1, and formaldehyde dismutase of Pseudomonas putida F61 revealed large differences in structural as well as catalytic properties, in spite of the fact that all are nicotinoproteins [enzymes which have bound NAD(P) as a cofactor]. It is concluded, therefore, that NDMA-ADH is a novel type of nicotinoprotein alcohol dehydrogenase.
Department of Microbiology and Enzymology, Delft University of Technology, The Netherlands.
P W
PW
Van Ophem
J
J
Van Beeumen
J A
JA
Duine
D039903:Actinobacteria / Q000201:enzymology* / Q000254:growth & development; D000429:Alcohol Oxidoreductases / Q000302:isolation & purification* / Q000378:metabolism; D000438:Alcohols / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002845:Chromatography; D002852:Chromatography, Ion Exchange; D017886:Durapatite; D006736:Horses; D006882:Hydroxyapatites; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D008969:Molecular Sequence Data; D009243:NAD / Q000032:analysis / Q000378:metabolism*; D009249:NADP / Q000032:analysis / Q000378:metabolism*; D012240:Rhodococcus / Q000201:enzymology; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D000438:Alcohols; D006882:Hydroxyapatites; D009243:NAD; D009249:NADP; D017886:Durapatite; D000429:Alcohol Oxidoreductases; C105563:alcohol - NDMA oxidoreductase