Purification of D-beta-hydroxybutyrate dehydrogenase from rat brain.

D-beta-Hydroxybutyrate dehydrogenase (BDH), a lipid-requiring enzyme, has been purified to homogeneity from rat brain using a new improved method. The purified rat brain BDH has a subunit molecular mass of 31 kilodaltons on sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The apoenzyme, i.e., the enzyme devoid of phospholipid, has no activity, but can be activated by phospholipid to a specific activity of 125 mumol/(min.mg). This is 625-fold greater than the activity in the mitochondrial fraction. The new purification procedure involves chromatography using a quaternary amine Sepharose resin followed by a sulphonate Sepharose resin, and eliminates the need for glass bead adsorption chromatography.