Phenacetin deacetylase activity in human liver microsomes: Distribution, kinetics, and chemical inhibition and stimulation

Microsomal and cytosolic phenacetin deacetylase activities were examined in human liver and kidneys. Kinetic properties of the activities were also st udied in human liver microsomes. Phenacetin deacetylase activity was predom inantly localized in the liver microsomal fraction. The specific activities of phenacetin deacetylation in liver cytosol and in kidney microsomes and cytosol were all less than 5% of that in liver microsomes. In human liver m icrosomes, Eadie-Hofstee plots for phenacetin deacetylation were monophasic , indicating a single-enzyme catalytic reaction. The Michaelis-Menten param eters, K-m and V-max, for the deacetylation were 4.7 mM and 5.54 nmol/min/m g of protein, respectively. The intrinsic clearance, calculated as V-max/K- m, was 1.18 ml/min/mg of protein. Although the organo-phosphate bis(4-nitro phenyl) phosphoric acid markedly inhibited the reaction in human liver micr osomes, the activity has a tolerance to the treatment of phenylmethylsulfon yl fluoride, a serine hydrolase inhibitor. Prazosin, a peripheral alpha(1)- adrenergic antagonist, noncompetitively inhibited the phenacetin deacetylat ion with a K-i value of 19.0 mu M. Flutamide, a nonsteroidal androgen recep tor antagonist, stimulated the activity by up to 349%. This increase was ac companied by a decrease in the K-m value and no change in the V-max value, resulting in an increase in the intrinsic clearance by up to 700% of the co ntrol. These results suggest that the phenacetin deacetylase localized in h uman liver microsomes has not only a catalytic site but also a negative and /or positive modulation site or sites.