The x-ray structure of D-amino acid oxidase at very high resolution identifies the chemical mechanism of flavin-dependent substrate dehydrogenation

Flavin is one of the most versatile redox cofactors in nature and is used b y many enzymes to perform a multitude of chemical reactions. D-Amino acid o xidase (DAAO). a member of the flavoprotein oxidase family, is regarded as a key enzyme for the understanding of the mechanism underlying flavin catal ysis. The very high-resolution structures of yeast DAAO complexed with D-al anine, D-trifluoroalanine, and L-lactate (1.20. 1.47, and 1.72 Angstrom) pr ovide strong evidence for hydride transfer as the mechanism of dehydrogenat ion. This is inconsistent with the alternative carbanion mechanism original ly favored for this type of enzymatic reaction. The step of hydride transfe r can proceed without involvement of amino acid functional groups. These st ructures, together with results from site-directed mutagenesis, point to or bital orientation/steering as the major factor in catalysis. A diatomic spe cies, proposed to be a peroxide, is found at the active center and on the R e-side of the flavin. These results are of general relevance for the mechan isms of flavoproteins and lead to the proposal of a common dehydrogenation mechanism for oxidases and dehydrogenases.