Substrate specificity of lysosomal cholesteryl ester hydrolase isolated from rat liver |
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Authors: | Rabbe Klemets Bo Lundberg |
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Affiliation: | (1) Wallac Biochemical Laboratory, P. O. Box 10, SF-20101 Turku 10, Finland;(2) Department of Biochemistry and Pharmacy, Abo Akademi, Abo, Finland |
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Abstract: | The effect of various physicochemical forms of substrate on the activity of acid cholesteryl ester hydrolase isolated from
rat liver lysosomes was studied. The amount of sodium taurocholate was varied in the substrate mixture which contained constant
amounts of egg phosphatidylcholine (PC) and cholesteryl oleate. The resulting substrate forms produced were PC vesicles, PC
vesicles with incorporated sodium taurocholate, mixed micelles, and mixed micelles together with free bile salt micelles.
Gradually increasing amounts of sodium taurocholate activated cholesteryl oleate hydrolysis until the molar sodium taurocholate/PC
ratio of ca. 0.6; thereafter hydrolytic activity decreased rapidly. The presence of sodium taurocholate micelles clearly inhibits
cholesteryl oleate hydrolysis. We therefore propose that the activation observed at low bile salt concentrations depends on
bile salt interaction with the substrate vehicle, whereas the inhibition observed at high bile salt concentrations depends
on sodium taurocholate interacting with the enzyme. When comparing different phospholipid components in the supersubstrate,
the enzyme activity was highest in the presence of dioleyl PC and decreased when present with dipalmitoyl PC and egg PC. Egg
lysoPC completely inhibited the enzyme activity. A net negative charge on the surface of the vesicle substrate increased cholesteryl
ester hydrolase activity while a net positive charge on the surface inhibited the enzyme activity. Only part of the product
inhibition of cholesteryl oleate hydrolase caused by Na-oleate was reversible when tested with bovine serum albumin present
in the incubation mixture. |
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