Purification and characterization of methanol dehydrogenase of Methylobacterium nodulans rhizosphere phytosymbionts

Methanol dehydrogenase (MDG) of the facultative methylotrophic phytosymbiont Methylobacterium nodulans has been purified for the first time to an electrophoretically homogeneous state and characterized. The native protein with a molecular mass of 70 kDa consists of large (60 kDa) and small (6 kDa) subunits. The purified protein displayed a specter identical to that of pyrochinolinchinon (PCC)-containing MDGs (pI 8.7, pH optimum in the range 9-10). The enzyme was inactive in the absence of ammonium or methylamine and exhibited a wide substrate specificity with regard to C1-C2 alcohols with the highest affinity to methanol (K(M) = 70 mM), but it did not oxidize benzyl and secondary alcohols. The apparent values of K(M) to primary alcohols increased with the length of the carbonic chain. The enzyme was characterized by a high stability level even in the absence of a substrate. An immobilized enzyme was used for amperometric methanol detection.