CHOLESTERYL ESTER LEVELS IN MAMMALIAN TISSUES

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The ability of intestinal carboxylester lipase (cholesterol esterase) to catalyze hydrolysis of substrates containing polyunsaturated and other fatty acids, requires the prior generation of lipolysis products by other lipases. Results obtained at interfaces comprised of either diacylglycerol or fatty acid product and a diacyl phosphatidylcholine suggest that the key to the activation of this enzyme activity is not the products per se, but their creation of areas of interface devoid of phosphatidylcholine. The all-or-none nature of the activation suggests that it depends critically on the lateral distribution of the phospholipid and substrate in the interfacial plane. On this basis we hypothesize a) that the critical transition will be observed in any interface which contains a lipid species to which carboxylester lipase cannot adsorb b) that the critical transition reflects changes in lateral distribution understandable in the context of percolation theory c) that the activation of carboxylester lipase by bile salts is also a consequence of percolation behavior and not the bilayer-to-micelle transition. These regulatory properties appear to be shared by hormone-sensitive lipase, an important intracellular lipase. Thus, we hypothesize d) that hormone-sensitive lipase is similarly regulated. To test these hypotheses we propose three specific aims 1) to correlate lipid chemical structure, carboxylester lipase adsorption and the occurrence of a critical transition in carboxylester lipase activity 2) to correlate the area normalized, lipid- composition dependence of the critical transition and rate constants for enzyme adsorption with i. lipid structure, ii. predictions of percolation theory and iii. parameters describing the non-ideality of lipid mixing, 3) to determine if bile salts can act as lipid spacers and, thereby, induce the critical transition in carboxylester lipase activity and 4) to determine how the phosphorylation state of hormone-sensitive lipase regulates its interfacial adsorption and catalysis. The achievement of the aims will not only test the hypotheses but will indicate how lipid head group and acyl interactions regulate lateral lipid distribution in interfaces. The critical activation phenomenon being investigated occurs with other lipases and phospholipases A and C. Because its regulation involves enzyme translocation from an aqueous to a surface phase and depends on the accumulation of acknowledged lipid messengers, like diacylglycerol and fatty acid, we believe the regulation being studied globally in this project reflects events occurring locally in cell membranes as a consequence of lipid-mediated signal transduction. Thus, the results from this project have a broader applicability to a poorly understood aspect of information transfer in cells.