UC Santa Barbara

The Group III Iron(II) Dependent Alcohol Dehydrogenases represent an understudied family of dehydrogenase enzymes. Although multiple members of the family have been cloned, expressed, and even crystallized and three-dimensional structure determined, very little work has gone into their chemical or kinetic mechanisms. Furthermore, it has been shown that certain members of the nudix hydrolase (nucleoside diphosphates linked to x) family of enzymes are capable of activating Group III dehydrogenases by two to tenfold. The method by which the nudix hydrolases cause activation is unknown. Using initial-rate and product inhibition studies in both non-activated and activated systems, the kinetic mechanism of one Group III Iron(II) Dependent Alcohol Dehydrogenase, gamma-hydroxybutyrate dehydrogenase (GHBDH) has been elucidated. Using mutational studies, the chemical mechanism of (GHBDH) was also studied. In GHBDH, the kinetic mechanism does not change upon activation by Nudix hydrolase; a proposed catalytic histidine does not appear to be necessary for the action of GHBDH. Finally, I have shown that in vitro studies of Group III ADHs may be missing a layer of metabolic control present in live cells.