Allylic or Benzylic Stabilization Is Essential for Catalysis by Bacterial Benzyl Alcohol Dehydrogenases

Benzyl alcohol dehydrogenase fromAcinetobacter calcoaceticus(AC-BADH) and TOL plasmid-encoded benzyl alcohol dehydrogenase fromPseudomonas putida(TOL-BADH) have previously been shown to oxidize a variety of aromatic alcohols but not aliphatic substrates. Here, we have expressed the genes for AC-BADH and TOL-BADH inEscherichia coli,purified the resulting over-expressed enzymes, and shown that each is an effective catalyst of both benzylic and allylic alcohol oxidation, but not of oxidation of nonallylic analogs. Enzyme specificity (kcat/Km) for both enzymes was higher with an aliphatic, allylic alcohol (3-methyl-2-buten-1-ol) than with benzyl alcohol. These results suggest that bacterial benzyl alcohol dehydrogenases use the resonance stabilization provided by allylic and benzylic alcohols to promote catalysis.