Functional sizes of hepatic enzymes of cholesteryl ester metabolism determined by radiation inactivation.
Cellular cholesteryl ester metabolism is regulated largely by the balance between intracellular esterification catalyzed by acyl-CoA:cholesterol acyltransferase and cholesteryl ester hydrolysis catalyzed by the cholesteryl ester hydrolases. The hydrolases include both cytosolic and membrane-associated activities; acidic and neutral activities have been described in both compartments. Esterification via the acyltransferase is membrane-associated. Neither the acyltransferase nor the membrane-associated hydrolases have been purified and characterized, and little is known about their genes. Thus, nothing is known about their sizes or structures. Radiation inactivation was used to determine the functional sizes in situ of acyl-coenzyme A:cholesterol acyltransferase, fatty acyl-CoA hydrolase, and acidic and neutral membrane-associated cholesteryl ester hydrolase activities. The functional M(r) +/- SD of the acyltransferase was 213 +/- 35 kD; for the acidic membrane-associated hydrolase, 48 +/- 2 kD; for the neutral membrane-associated hydrolase, 94 +/- 15 kD; and for the fatty acyl-CoA hydrolase, 65 +/- 15 kD. Monoexponential curves were observed in all cases using radiation exposures that inactivated enzyme activities to < or = 10% of control values. Substrate specificity and inhibition studies suggested that the active sites of the acyltransferase and fatty acyl-CoA hydrolase were different, supporting the concept that the hydrolase is not part of the functional unit required for cholesterol esterification.
Erickson
S K
SK
Department of Medicine, University of California, San Francisco.
Lear
S R
SR
McCreery
M J
MJ
D000818:Animals; D002788:Cholesterol Esters / Q000378:metabolism*; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology* / Q000528:radiation effects; D008970:Molecular Weight; D010170:Palmitoyl-CoA Hydrolase / Q000528:radiation effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D002787:Sterol Esterase / Q000528:radiation effects; D002785:Sterol O-Acyltransferase / Q000528:radiation effects
D002788:Cholesterol Esters; D002785:Sterol O-Acyltransferase; D002787:Sterol Esterase; D010170:Palmitoyl-CoA Hydrolase