Adenosylcobalamin-mediated methyl transfer by toluate cis-dihydrodiol dehydrogenase of the TOL plasmid pWWO

We identified and characterized a methyl transfer activity of the toluate c is-dihydrodiol (4-methyl-3,5-cyclohexadiene-cis-1,2-diol-1-carboxylic acid) dehydrogenase of the TOL plasmid pWW0 towards toluene cis-dihydrodiol (3-m ethyl-4,5-cyclohexadiene-cis-1,2-diol). When the purified enzyme from the r ecombinant Escherichia coli containing the xylL gene was incubated with tol uene cis-dihydrodiol in the presence of NAD(+), the end products differed d epending on the presence of adenosylcobalamin (coenzyme B-12). The enzyme y ielded catechol in the presence of adenosylcobalamin, while it gave 3-methy lcatechol in the absence of the cofactor. Adenosyl-cobalamin was transforme d to methylcobalamin as a result of the enzyme reaction, which indicates th at the methyl group of the substrate was transferred to adenosylcobalamin. Other derivatives of the cobalamin such as aquo (hydroxy)- and cyanocobalam in did not mediate the methyl transfer reaction. The dehydrogenation and me thyl transfer reactions were assumed to occur concomitantly, and the methyl transfer reaction seemed to depend on the dehydrogenation. To our knowledg e, the enzyme is the first dehydrogenase that shows a methyl transfer activ ity as well.