CHARACTERISTICS OF A PURIFIED DOG HEPATIC-MICROSOMAL N,O-ACYLTRANSFERASE

Dog liver microsomes have at least three different enzymes that are ca pable of the deacylation of amides, N-arylhydroxamic acids and carboxy lesters, the acyltransfer of N-arylhydroxamic acids and the N-acetylat ion of arylamines. As judged by SDS - PAGE stained with silver nitrate , one of these enzymes was purified to homogeneity by sequential treat ment with Triton X-100, ion-exchange column chromatography, gel filtra tion and chromatofocusing. The protein was a glycoprotein trimer with a subunit weight of similar to 60 kDa. It showed microheterogeneity on analytical isoelectric focusing (IEF) in polyacrylamide with pls of 5 .4-5.6. Following digestion with endoglycosidase H, its subunit weight was reduced to similar to 58 kDa, and it appeared to be homogeneous o n IEF with a pi of similar to 5.6. A monoclonal antibody prepared agai nst this enzyme also reacted with the pi 6.0 carboxylesterase of rat l iver microsomes, but did not react with the other two dog hepatic acyl transferases. Conversely, a polyclonal antibody raised against the rat esterase reacted with the dog enzyme. The N-terminal sequence of the enzyme was Y-P-S-GP-P-V-V-D-T-V-Q-G-K-V-, which was homologous to the form 1 carboxylesterase of rabbit liver and the pl 6.0 carboxylesteras e of rat liver. Immunohistochemical analyses showed the presence of th is enzyme in the epithelium of dog liver and urinary bladder, human li ver and rat liver, esophagus, forestomach, glandular stomach, small an d large intestines, renal tubules, trachea and prostate and alveolar c ells of lung. Since this enzyme is present in the urothelium, it may b e important for the activation of urinary metabolites of carcinogenic arylamines for the initiation of bladder carcinogenesis in the dog.