Target size of D-beta-hydroxybutyrate dehydrogenase. Functional and structural molecular weight based on radiation inactivation.
D-beta-Hydroxybutyrate dehydrogenase is a lipid-requiring enzyme which is localized on the inner face of the mitochondrial inner membrane. The apoenzyme has been purified to homogeneity from beef heart; it is devoid of lipid and inactive. It can be functionally reconstituted with lecithin or phospholipid mixtures containing lecithin. The active form of the enzyme is the enzyme-phospholipid complex. Classical target analysis of radiation-inactivation data has now been used to determine the molecular size of the enzyme both in the native membrane (submitochondrial vesicles) and in the reconstituted enzyme inserted into phospholipid vesicles containing lecithin. For both forms of the enzyme, we find the same molecular size, approximately 110,00 daltons. This size is consistent with a tetramer. Radiation results in fragmentation of the polypeptide and the destruction of the polypeptide correlates with loss of enzymic function. A similar size is obtained when purified D-beta-hydroxybutyrate dehydrogenase is inserted into a nonactivating mixture of phospholipid (i.e. in the absence of lecithin). We conclude that: 1) the native enzyme in submitochondrial vesicles and the purified active enzyme in phospholipid vesicles are the same size, approximating a tetramer; 2) radiation of D-beta-hydroxybutyrate dehydrogenase results in loss of activity and fragmentation of the polypeptide; and 3) the role of lecithin in activation of D-beta-hydroxybutyrate dehydrogenase is unrelated to determining oligomeric size of the enzymes since both active and nonactive forms exhibit the same structural size.
D000818:Animals; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D006884:Hydroxybutyrate Dehydrogenase / Q000378:metabolism* / Q000528:radiation effects; D008929:Mitochondria, Heart / Q000201:enzymology; D008970:Molecular Weight
D006884:Hydroxybutyrate Dehydrogenase