Purification and characterization of rhesus monkey liver amido hydrolases and their roles in the metabolic polymorphism for E6123, a platelet-activating factor receptor antagonist.
We previously showed that a polymorphism for E6123 [(S)-(+)-6- (2-chlorophenyl)-3-cyclopropanecarbonyl-8,11-dimethyl-2,3,4,5- tetrahydro-8H-pyrido[4',3':4,5]thieno[3,2-f][1,2,4]triazolo[4,3-a] [1,4]diazepine] metabolism exists only in rhesus monkeys. In the present study, we purified, from rhesus monkey hepatic microsomes, three amido hydrolases that are involved in the metabolic polymorphism. Two forms of amido hydrolase from an extensive metabolizer and one from a poor metabolizer were purified by Q-Sepharose Fast Flow, Red A-agarose, octylamino-Sepharose 4B, and hydroxyapatite-Ultrogel chromatography, after solubilization with Lubrol. The three purified enzymes had the same molecular mass (47 kDa), and their amino-terminal amino acid sequences were identical. The enzymes were different from various known carboxylesterases in terms of substrate specificity, molecular mass, and amino-terminal amino acid sequence. They resembled arylacetamide deacetylase from human hepatic microsomes with respect to molecular mass and amino-terminal amino acid sequence. The KM values of the high and low affinity enzymes in the extensive metabolizer and the sole enzyme in the poor metabolizer were 37.6, 73.0, and 76.5 microM, respectively. The Vmax values were 3312.4, 504.8, and 427.9 pmol/min/mg of protein, respectively. The high affinity enzyme in extensive metabolizer appears to be quite distinct, whereas the low affinity enzyme in extensive metabolizer in similar or identical to the sole enzyme in poor metabolizer. Thus, the metabolic polymorphism in rhesus monkey may depend upon the existence of the high affinity enzyme in extensive metabolizer.
Kusano
K
K
Tsukuba Research Laboratories, Eisai Co., Ltd., Ibaraki, Japan.
Seko
T
T
Tanaka
S
S
Shikata
Y
Y
Ando
T
T
Ida
S
S
Hosokawa
M
M
Satoh
T
T
Yuzuriha
T
T
Horie
T
T
D000581:Amidohydrolases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001381:Azepines / Q000494:pharmacology*; D002853:Chromatography, Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D008099:Liver / Q000201:enzymology*; D008253:Macaca mulatta; D008297:Male; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010980:Platelet Membrane Glycoproteins / Q000037:antagonists & inhibitors*; D011110:Polymorphism, Genetic; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D014230:Triazoles / Q000494:pharmacology*
D001381:Azepines; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D014230:Triazoles; C039351:platelet activating factor receptor; C068033:E 6123; D000581:Amidohydrolases