title: Overexpression, purification and characterization of Mycobacterium bovis BCG alcohol dehydrogenase.
abstract: A previous study of the effect of zinc deprivation on Mycobacterium bovis BCG pointed out the potential importance of an alcohol dehydrogenase for maintaining the hydrophobic character of the cell envelope. In this report, the effect of the overexpression of the M. bovis BCG alcohol dehydrogenase (ADH) in Mycobacterium smegmatis and M. bovis BCG is described. The purification 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. The best kcat/Km values were found with benzaldehyde > 3-methoxybenzaldehyde > octanal > coniferaldehyde. A phylogenetic analysis clearly revealed that 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 alcohol dehydrogenases, the plant cinnamyl alcohol dehydrogenases (CADs). Comparison of the kinetic properties of our ADH with some related class C enzymes indicated that the mycobacterial enzyme substrate profile resembled that of the CADs involved in plant defence rather than those implicated in lignification. A possible role for the M. bovis BCG ADH in the biosynthesis of the lipids composing the mycobacterial cell envelope is proposed.
authors.affiliation: Institut Pasteur de Bruxelles, Belgium.
authors.forename: J M
authors.initials: JM
authors.lastname: Wilkin
authors.forename: K
authors.initials: K
authors.lastname: Soetaert
authors.forename: M
authors.initials: M
authors.lastname: Stélandre
authors.forename: P
authors.initials: P
authors.lastname: Buyssens
authors.forename: G
authors.initials: G
authors.lastname: Castillo
authors.forename: V
authors.initials: V
authors.lastname: Demoulin
authors.forename: G
authors.initials: G
authors.lastname: Bottu
authors.forename: M A
authors.initials: MA
authors.lastname: Laneelle
authors.forename: M
authors.initials: M
authors.lastname: Daffe
authors.forename: J
authors.initials: J
authors.lastname: De Bruyn
mesh_terms: D000426:Alcohol Dehydrogenase / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D001483:Base Sequence; D017931:DNA Primers; D007700:Kinetics; D009163:Mycobacterium bovis / Q000201:enzymology*; D010641:Phenotype; D010802:Phylogeny
chemical_list: D017931:DNA Primers; D000426:Alcohol Dehydrogenase