Open AccessProperties of an NAD(H)‐Containing Methanol Dehydrogenase and its Activator Protein from Bacillus methanolicus
Author(s) -
Arfman Nico,
Hektor Harm J.,
Bystrykh Leonid V.,
Govorukhitalya I.,
Dijkhuizen Lubbert,
Frank Johannes
Publication year - 1997
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1997.00426.x
Subject(s) - nad+ kinase , cofactor , methanol dehydrogenase , enzyme , chemistry , glycerol 3 phosphate dehydrogenase , biochemistry , formate dehydrogenase , oxidoreductase , stereochemistry , biology
Oxidation of C 1 –C 4 primary alcohols in thermotolerant Bacillus methanolicus strains is catalyzed by an NAD‐dependent methanol dehydrogenase (MDH), composed of ten identical 43000‐M r subunits. Each MDH subunit contains a tightly, but non‐covalently, bound NAD(H) molecule, in addition to 1 Zn 2+ and 1–2 Mg 2+ ions. The NAD(H) cofactor is oxidized and reduced by formaldehyde and methanol, respectively, while it remains bound to the enzyme. Incubation of MDH with methanol and exogenous NAD (coenzyme) results in reduction of this NAD coenzyme. Both NAD species are not exchanged during catalysis. NAD thus plays two different and important roles in the MDH‐catalyzed reaction, with the bound NAD cofactor acting as primary electron acceptor and the NAD coenzyme being responsible for reoxidation of the reduced cofactor. MDH obeys a ping‐pong type reaction mechanism, which is consistent with such a temporary parking of reducing equivalents at the MDH‐bound cofactor. Spectral studies show that, in the presence of exogenous NAD and Mg 2+ ions, MDH interacts with a previously identified 50000‐M r activator protein. The activator protein appears to facilitate the oxidation of the reduced NADH cofactor of MDH, which results in a strongly increased turnover rate of MDH.