Overexpression, purification and characterization of Mycobacterium bovis BCG alcohol dehydrogenase

A previous study of the effect of zinc deprivation on Mycobacterium bovis B CG pointed out the potential importance of an alcohol dehydrogenase for mai ntaining the hydrophobic character of the cell envelope. In this report, th e effect of the overexpression of the M. bovis BCG alcohol dehydrogenase (A DH) in Mycobacterium smegmatis and M. bovis BCG is described. The purificat ion of the enzyme was performed to apparent homogeneity from overexpressing M. bovis BCG cells and its kinetic parameters were determined. The enzyme showed a strong preference for both aromatic and aliphatic aldehydes while the corresponding alcohols were processed 100-1000-fold less efficiently. T he best k(cat)/K-m values were found with benzaldehyde > 3-methoxybenzaldeh yde > octanal > coniferaldehyde. A phylogenetic analysis clearly revealed t hat the M. bovis BCG ADH together with the ADHs from Bacillus subtilis and Helicobacter pylori formed a sister group of the class C medium-chain alcoh ol dehydrogenases, the plant cinnamyl alcohol dehydrogenases (CADs). Compar ison of the kinetic properties of our ADH with some related class C enzymes indicated that the mycobacterial enzyme substrate profile resembled that o f the CADs involved in plant defence rather than those implicated in lignif ication. A possible role for the M. bovis BCG ADH in the biosynthesis of th e lipids composing the mycobacterial cell envelope is proposed.