Purification and characterization of rat liver enzyme catalyzing stereoselective reduction of acetylpyridines.
Acetylpyridines (1-3) are known as aroma components of foods, perfumes, and smoking suppressants, showing several biological activities and constituting part of the structure of some important biologically active compounds. We purified and characterized an enzyme that catalyzes the stereoselective reduction of acetylpyridines so that we could clarify its function. The enzyme participating in the reductive metabolism of 4-acetylpyridine (1) in the rat liver was purified by successively applying ammonium sulfate fractionation, anion-exchange, gel filtration, and affinity chromatography, and it was definitively identified as 3alpha-HSD. It preferentially reduced acetylpyridines (1-3) and acetophenone (7) to their corresponding (S)-alcohols, with high enantioselectivity. Kinetic analyses of the compounds were performed, and the V(max)/K(m) values decreased in the order of 4-, 2-, and 3-acetylpyridine (1, 3, 2), while acetophenone (7) showed almost the same value as 3-acetylpyridine (2). These results suggested that the reduction of the substrates by 3alpha-HSD is affected by the nitrogen atom in the aromatic ring.
Uwai
Koji
K
Department of Pharmaceutics, Tohoku Pharmaceutical University, Sendai, Japan. uwai@tohoku-pharm.ac.jp
Konno
Noboru
N
Kitamura
Shigeyuki
S
Ohta
Shigeru
S
Takeshita
Mitsuhiro
M
D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific) / Q000737:chemistry* / Q000302:isolation & purification*; D000595:Amino Acid Sequence; D000818:Animals; D002384:Catalysis; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010084:Oxidation-Reduction; D011725:Pyridines / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid; D013237:Stereoisomerism; D013329:Structure-Activity Relationship
D011725:Pyridines; C010835:3-acetylpyridine; D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific); C055538:4-acetylpyridine