Characteristics of a purified dog hepatic microsomal N,O-acyltransferase.
Dog liver microsomes have at least three different enzymes that are capable of the deacylation of amides, N-arylhydroxamic acids and carboxylesters, the acyltransfer of N-arylhydroxamic acids and the N-acetylation of arylamines. As judged by SDS-PAGE stained with silver nitrate, one of these enzymes was purified to homogeneity by sequential treatment with Triton X-100, ion-exchange column chromatography, gel filtration and chromatofocusing. The protein was a glycoprotein trimer with a subunit weight of approximately 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 approximately 58 kDa, and it appeared to be homogeneous on IEF with a pl of approximately 5.6. A monoclonal antibody prepared against this enzyme also reacted with the pl 6.0 carboxylesterase of rat liver microsomes, but did not react with the other two dog hepatic acyltransferases. 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-L-P-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 carboxylesterase of rat liver. Immunohistochemical analyses showed the presence of this enzyme in the epithelium of dog liver and urinary bladder, human liver and rat liver, esophagus, forestomach, glandular stomach, small and large intestines, renal tubules, trachea and prostate and alveolar cells of lung. Since this enzyme is present in the urothelium, it may be important for the activation of urinary metabolites of carcinogenic arylamines for the initiation of bladder carcinogenesis in the dog.
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Department of Chemical Carcinogenesis, Michigan Cancer Foundation, Detroit 48201.
Zukowski
K
K
Land
S J
SJ
King
C M
CM
Martin
B M
BM
Pohl
L R
LR
Wang
C Y
CY
D000217:Acyltransferases / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001711:Biotransformation; D002265:Carboxylic Ester Hydrolases / Q000737:chemistry / Q000378:metabolism; D004285:Dogs; D007526:Isoelectric Point; D008562:Membrane Glycoproteins / Q000302:isolation & purification; D008862:Microsomes, Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D001743:Urinary Bladder / Q000201:enzymology
D008562:Membrane Glycoproteins; D000217:Acyltransferases; D002265:Carboxylic Ester Hydrolases