title: Solubilization, partial purification and characterization of rat liver microsomal diacylglycerol acyltransferase.
abstract: The diacylglycerol acyltransferase (EC 2.3.1.20) activity of rat liver microsomes was solubilized with deoxycholate, cholate, and Triton X-100. The cholate-solubilized activity proved to be more stable than the deoxycholate-solubilized activity. Cholate caused less interference with the diacylglycerol acyltransferase assay than Triton X-100. The cholate-solubilized activity was purified 3- to 4-fold by Sepharose 4B chromatography, and could be separated from most of the cholate-solubilized protein by centrifugation in a 10--20% sucrose gradient. The purification led to an overall purification of 9-fold with a recovery of 80%. The gradient fractions were analyzed for protein, RNA, and phospholipid content as well as for several enzyme activities of glycerolipid biosynthesis. The partially purified fractions were delipidated and activity was stimulated 5-fold by the addition of sonicated microsomal phospholipids. The soluble, gradient-puridied diacylglycerol acyltransferase activity was strongly dependent on added magnesium.
mesh_terms: D000217:Acyltransferases / Q000302:isolation & purification* / Q000378:metabolism; D000818:Animals; D002499:Centrifugation, Density Gradient; D002793:Cholic Acids; D002847:Chromatography, Agarose; D003840:Deoxycholic Acid; D051048:Diacylglycerol O-Acyltransferase; D005260:Female; D008274:Magnesium / Q000494:pharmacology; D008862:Microsomes, Liver / Q000201:enzymology*; D011092:Polyethylene Glycols; D051381:Rats; D012995:Solubility
chemical_list: D002793:Cholic Acids; D003840:Deoxycholic Acid; D011092:Polyethylene Glycols; D000217:Acyltransferases; D051048:Diacylglycerol O-Acyltransferase; D008274:Magnesium