Abstract
The properties of ES46.5K, an esterase from mouse hepatic microsomes, were compared with those of carboxylesterases from rabbit and porcine liver. The inhibitory profile with a serine hydrolase inhibitor (bis-p-nitrophenylphosphate) and detergents (sodium dodecylsulfate, Emulgen 911) was different between ES46.5K and the carboxylesterases. Bis-p-nitrophenylphosphate (0.1 mM) markedly inhibited the catalytic activity of the carboxylesterases but not that of ES46.5K. Emulgen 911 (0.05-0.25%) inhibited the catalytic activity of the carboxylesterases, whereas the detergent conversely stimulated that of ES46.5K by 150%. The two carboxylesterases catalyzed the hydrolysis of acetate esters of tetrahydrocannabinol (THC) analogs with different side chain lengths (C1-C5), although ES46.5K showed marginal activity only against the acetate of D8-tetrahydrocannabiorcol, a Me side chain deriv. of D8-THC. ES46.5K hydrolyzed cannabinoid esters stereospecifically and regioselectively. The esterase hydrolyzed 8a-acetoxy-D9-tetrahydrocannabinol (8a-acetoxy-D9-THC, 5.62 nmol/min/mg protein), while the enzyme did not hydrolyze 8b-acetoxy-D9-THC, 7a-acetoxy-, and 7b-acetoxy-D8-THC at all. In contrast, the carboxylesterases from rabbit and porcine liver hydrolyzed 8b-acetoxy-D9-THC efficiently but not 8a-acetoxy-D9-THC. ES46.5K hydrolyzed side chain acetoxy derivs. of D8-THC at the 3'- and 4'-positions, and a Me ester of 5'-nor-D8-THC-4'-oic acid. The enzyme, however, could not hydrolyze Me esters of D8- and D9-THC-11-oic acid, while both carboxylesterases hydrolyzed side chain acetoxy derivs. of D8-THC and three Me esters of THC-oic acids. These differences in stereospecificity and regioselectivity between ES46.5K and carboxylesterases suggest that the configurations of important amino acids for the catalytic activities of these enzymes are different from each other.
