PHYSICO-CHEMICAL REGULATN OF ENZYMES IN LIPID METABOLISM

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The goal of this project is to determine the mechanisms by which the phospholipase A2 and colipase-dependent lipase from pancreas are regulated at lipid-water phase boundaries. It is hypothesized that a major factor in this regulation is the lateral organization of lipids in the plane of the lipid-water interface. Such regulation is expressed both at the level of enzyme partitioning to the interface from aqueous solution and at the level of catalysis within the interfacial plane. Emphasis is on understanding the roles of lipid composition, packing, surface potential and the protein cofactor, colipase, in the initiation and maintenance of phosphatidylcholine and glyceride hydrolysis, i.e., (phospho) lipolysis. To identify the extent to which organization influences (phospho)lipolysis, enzyme adsorption to and catalysis in model phospholipid-glyceride interfaces will be measured. This will include both initial rate measurements and measurement of the extent of adsorption and catalysis. The interfaces will be monolayers and bilayers which are highly controlled with respect to lipid composition, packing density and electrical properties. By correcting results for the contribution of the usual intensive variables of enzyme and substrate concentrations, organization-dependent effects will be revealed. When examined on a lipid composition basis, these corrected data will reveal the composition at which the postulated critical transition occurs and how relative lipid cluster size changes as the critical composition is approached. In related experiments, average cluster size will be measured. These studies address the regulation of interfacial reactions which occur not only in the intestine but in all tissues. Such reactions are involved in inflammatory processes, lipid transport and signal transduction. Their control, or lack of it, contributes to the pathology of diseases such as atherosclerosis and arthritis.