Tetrazolium-dye-linked alcohol dehydrogenase of the methylotrophic actinomycete Amycolatopsis methanolica is a three-component complex.
Tetrazolium-dye-linked alcohol dehydrogenase (TD-ADH) of Amycolatopsis methanolica could be resolved into three protein components, which have been purified. Each of the components has the ability to reconstitute TD-ADH activity when combined with the other two. Component 1 is identical to the previously characterized methanol:N,N'-dimethyl-4-nitrosoaniline oxidoreductase (MNO), a decameric protein with 50-kDa subunits, each carrying a tightly bound NADPH. Component 2 is a high molecular mass (> 640 kDa) protein with subunits of 44 kDa and 72 kDa, and which possesses a low tetrazolium-dye-linked NADH dehydrogenase activity. The protein contains a yellow chromophore of unknown identity. Component 3 is a low molecular mass (15 kDa) protein containing a 5'-deazaflavin and at least one other low-molecular-mass compound with properties similar, but not identical, to those of nicotinamide coenzymes. The results suggest that alcohol oxidation by the TD-ADH complex is carried out by component 1 (MNO), after which transfer of the reducing equivalents (mediated by component 3) occurs to component 2, which (in vitro) is linked to the tetrazolium dye. Fractionation of A. methanolica extracts showed that most of the 5'-deazaflavin was present in component 3. Other gram-positive bacteria having a TD-ADH complex also produced 5'-deazaflavin. It is concluded that oxidation of primary aliphatic alcohols by A. methanolica, and probably also by other gram-positive bacteria containing MNO or TD-ADH, proceeds via TD-ADH. The likeliness of 5'-deazaflavin participation in this process is discussed.
Bystrykh
L V
LV
Department of Microbiology and Enzymology, Delft University of Technology, The Netherlands.
Govorukhina
N I
NI
Dijkhuizen
L
L
Duine
J A
JA
D039903:Actinobacteria / Q000201:enzymology*; D000426:Alcohol Dehydrogenase / Q000737:chemistry*; D000429:Alcohol Oxidoreductases / Q000378:metabolism; D005415:Flavins / Q000737:chemistry; D013778:Tetrazolium Salts / Q000737:chemistry; D013844:Thiazoles / Q000737:chemistry
D005415:Flavins; D013778:Tetrazolium Salts; D013844:Thiazoles; D000429:Alcohol Oxidoreductases; D000426:Alcohol Dehydrogenase; C019641:alcohol dehydrogenase (acceptor); C022616:thiazolyl blue