Purification and some properties of a monoacylglycerol-hydrolyzing enzyme of rat adipose tissue.
A monoacylglycerol-hydrolyzing enzyme has been purified 2500-fold from rat adipose tissue. The key step was the solubilization of the enzyme, presumably as an enzyme-detergent complex, by sonication with a nonionic polyoxyethylene alcohol detergent. The purification was achieved by ion exchange and gel chromatography, and isoelectric focusing, in the presence of detergent. By sodium dodecyl sulfate gel electrophoresis the enzyme protein was more than 85% pure. This method indicated a minimum molecular weight of 32,900. The preliminary amino acid composition, excluding tryptophan, could best be fitted with a value of 31,800. The purified enzyme had a pI of 7.2, an estimated Stokes radius of 39 A by gel chromatography and a pH optimum of 8.0. Enzyme stability was highly dependent on presence of detergent and free sulfhydryl groups. The enzyme was responsible for the main monoacylglycerol- but only a small part of the p-nitrophenylacetate-hydrolyzing activity of crude adipose tissue extracts and hydrolyzed 1(3)- and 2-monooleoylglycerol at equal rates. Under the assay conditions used it did not catalyze the hydrolysis of emulsified trioleoylglycerol, micellar or emulsified dioleoylglycerol, emulsified cholesterol oleate or micellar lysophosphatidylcholine. It is possible that the enzyme may be a specific monoacylglycerol hydrolase.
D000273:Adipose Tissue / Q000201:enzymology*; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002265:Carboxylic Ester Hydrolases / Q000302:isolation & purification*; D002729:Chloromercuribenzoates / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D007700:Kinetics; D008297:Male; D008970:Molecular Weight; D008994:Monoacylglycerol Lipases / Q000302:isolation & purification* / Q000378:metabolism; D011487:Protein Conformation; D051381:Rats; D013501:Surface-Active Agents
D000596:Amino Acids; D002729:Chloromercuribenzoates; D013501:Surface-Active Agents; D002265:Carboxylic Ester Hydrolases; D008994:Monoacylglycerol Lipases; D004229:Dithiothreitol