Effects of inhibitors of N-linked oligosaccharide processing on the secretion, stability, and activity of lecithin:cholesterol acyltransferase.
The structure and function of the carbohydrate moiety of human lecithin:cholesterol acyltransferase (LCAT) were determined by using several glycosidases in reaction with the isolated plasma protein or by using specific inhibitors of glycoprotein assembly with cultured cells secreting LCAT activity. Analysis of the plasma enzyme indicated that almost all of the large carbohydrate moiety of LCAT (approximately 25% w/w) was N-linked with part of the high-mannose and part of the complex type. This analysis was confirmed with metabolic inhibitors of carbohydrate processing by using CHO cells stably transfected with the human LCAT gene. Inhibitors of the subsequent processing of the N-linked high-mannose chains formed by glucosidase activity were without effect on either the secretion rate or the catalytic activity of LCAT. The inhibition of catalytic activity by glucosidase inhibitors applied to both the phospholipase and the acyltransferase activities of LCAT. The reduction of the LCAT catalytic rate by terminal glycosidase inhibitors was without effect on apparent Km and did not affect enzyme stability. These data indicate an unusual specific role for high-mannose carbohydrates in the catalytic mechanism of LCAT.
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Cardiovascular Research Institute, University of California Medical Center, San Francisco 94143.
Fielding
C J
CJ
D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D002460:Cell Line; D004795:Enzyme Stability; D006026:Glycoside Hydrolases / Q000037:antagonists & inhibitors*; D007212:Indolizines / Q000494:pharmacology; D007700:Kinetics; D008361:Mannosidases / Q000037:antagonists & inhibitors; D009844:Oligosaccharides / Q000378:metabolism*; D007862:Phosphatidylcholine-Sterol O-Acyltransferase / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects*; D017026:Swainsonine; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics; D013816:Thermodynamics; D014162:Transfection; D014415:Tunicamycin / Q000494:pharmacology*; D054836:Indolizidines
D000470:Alkaloids; D000900:Anti-Bacterial Agents; D006026:Glycoside Hydrolases; D007212:Indolizines; D008361:Mannosidases; D009844:Oligosaccharides; D007862:Phosphatidylcholine-Sterol O-Acyltransferase; D017026:Swainsonine; D013762:Tetrahydrofolate Dehydrogenase; D014415:Tunicamycin; C037806:castanospermine; D054836:Indolizidines