Enzymic and structural studies on Drosophila alcohol dehydrogenase and other short-chain dehydrogenases/reductases

Enzymic and structural studies on Drosophila alcohol dehydrogenases and oth er short-chain dehydrogenases/reductases (SDRs) are presented. Like alcohol dehydrogenases from other Drosophila species, the enzyme from D, simulans is more active on secondary than on primary alcohols, although ethanol is i ts only known physiological substrate. Several secondary alcohols were used to determine the kinetic parameters k(cat) and K,, The results of these ex periments indicate that the substrate-binding region of the enzyme allows o ptimal binding of a short ethyl side-chain in a small binding pocket, and o f a propyl or butyl side-chain in large binding pocket, with stereospecific ity for R(-) alcohols. At a high concentration of R(-) alcohols substrate a ctivation occurs. The k(cat) and K-m values determined under these conditio ns are about two-fold, and two orders of magnitude , respectively, higher t han those at low substrate concentrations. Sequence alignment of several SDRs of known, and unknown three-dimensional structures, indicate the presence of several conserved residues in addition to those involved in the catalyzed reactions. Structural roles of these co nserved residues could be derived from observations made on superpositioned structures of several SDRs with known structures. Several residues are con served in tetrameric SDRs, but not in dimeric ones. Two halohydrin-halide-l yases show significant homology with SDRs in the catalytic domains of these enzymes, but they do not have the structural features required for binding NAD(+). Probably these lyases descend from an SDR, which has lost the capa bility to bind NAD(+), but the enzyme reaction mechanisms may still be simi lar.