Purification and characterization of dimeric dihydrodiol dehydrogenase from dog liver.
High NADP(+)-linked dihydrodiol dehydrogenase activity was detected in dog liver cytosol, from which a dimeric enzyme composed of M(r) 39,000 subunits was purified to homogeneity. The enzyme oxidized trans-cyclohexanediol, and trans-dihydrodiols of benzene and naphthalene, the [1R,2R]-isomers of which were selectively oxidized. In the reverse reaction in the presence of NADPH as a coenzyme, the enzyme reduced alpha-dicarbonyl compounds, such as methylglyoxal, 3-deoxyglucosone, and diacetyl, and some compounds with a carbonyl group, such as glyceraldehyde, lactaldehyde, and acetoin. 4-Hydroxyphenylketones and ascorbates inhibited the enzyme. The results of steady-state kinetic analyses indicated that the reaction proceeds through an ordered bi bi mechanism with the coenzyme binding to the free enzyme, and suggested that the inhibitors bind to the enzyme-NADP+ binary complex. The dimeric enzyme was detected in liver and kidney of dog, and was immunochemically similar to the dimeric enzymes from monkey kidney, rabbit lens, and pig liver. The sequences (total 127 amino acid residues) of eight peptides derived on enzymatic digestion of the dog liver enzyme did not show significant similarity with the primary structures of members of the aldo-keto reductase and short chain dehydrogenase superfamilies, which include monomeric dihydrodiol dehydrogenases and carbonyl reductase, respectively.
Sato
K
K
Biochemistry Laboratory, Gifu Pharmaceutical University.
Nakanishi
M
M
Deyashiki
Y
Y
Hara
A
A
Matsuura
K
K
Ohya
I
I
D000818:Animals; D001704:Biopolymers; D004285:Dogs / Q000378:metabolism*; D000882:Haplorhini; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008297:Male; D008970:Molecular Weight; D009928:Organ Specificity / Q000502:physiology; D010088:Oxidoreductases / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D011817:Rabbits; D013045:Species Specificity; D013379:Substrate Specificity; D013552:Swine
D001704:Biopolymers; D010088:Oxidoreductases; C048338:trans-1,2-dihydrobenzene-1,2-diol dehydrogenase