Cholesterol ester metabolism in the brain: Properties and subcellular distribution of cholesterol-esterifying enzymes and cholesterol ester hydrolases in adult rat brain

Abstract

1. 1. The properties and subcellular distribution of the enzymes directly related to the formation and degradation of cholesterol esters were investigated in the young adult rat brain. They were a cholesterol-esterifying enzyme, lecithin-cholesterol acyltransferase, and two distinct cholesterol ester hydrolases. 2. 2. The cholesterol-esterifying enzyme incorporated exogenous free [1- 14C]-oleic acid into cholesterol ester. The pH optimum was 5.5–5.6. Exogenous ATP or CoA did not stimulate the activity, and the presence of an excess amount of unlabelled oleyl-CoA did not diminish the incorporation of the radioactive oleic acid. Various bile acids and Tween 20 were inhibitory, particularly at higher concentrations. Under the optimum conditions, the total activity of the brain was approximately 40 μg cholesterol esterified per h per g. Nearly half of the activity was found in the crude mitochondrial fraction. Purified myelin contained no activity. 3. 3. All attempts failed to demonstrate the activity of lecithin-cholesterol acyltranferase in the adult rat brain, using the specific substrate, lecithin labelled at the β-position with [1- 14C]oleic acid. The contribution of this enzyme to the formation of cholesterol ester in normal adult rat brain appears to be, if not nil, negligible. 4. 4. Two distinct cholesterol ester hydrolases were present. One had the pH optimum of 4.2, was activated by deoxycholate, taurocholate, and cholate, and moderately inhibited by Tween 20. Under the optimum condition in the presence of taurocholate, the total activity of this enzyme was approximately 30μg of cholesteryl oleate hydrolyzed per h per g. This enzyme was primarily localized in the crude mitochondrial fraction, and the subfractionation of the crude mitochondrial fraction suggested that this enzyme might be localized in mitochondria, rather than in lysosomes. Purified myelin was devoid of the pH 4.2 cholesterol ester hydrolase. 5. 5. The other cholesterol ester hydrolase had the pH optimum of 6.6. It was strongly inhibited by deoxycholate and activated by taurocholate. Cholate activated the enzyme at a low concentration, but was moderately inhibitory at a higher concentration. At the optimum condition with added taurocholate, the total activity of the brain was about 170 μg cholesteryl oleate hydrolyzed per h per g. The bulk of this enzyme appeared to be localized in microsomes. However, the highly purified myelin fraction was also enriched with this enzyme by more than 3-fold, compared to the starting homogenate, and it retained 20% of the total activity of the pH 6.6 cholesterol ester hydrolase in the homogenate.