Phospholipid transfer (exchange) proteins catalyze the translocation of intact phospholipid molecules between natural and artificial membranes. Present in a number of bovine tissues, but chiefly cerebral cortex, is one of these proteins which exhibits specificity for phosphatidylinositol and, to a lesser extent, phosphatidylcholine. Under appropriate conditions, net transfer of individual phospholipids or equivalent directional exchange of total phospholipid can be shown. A critical step in the catalytic process is the formation of a complex between the transfer protein and a membrane surface, at which point protein-bound and membrane-bound phospholipids are exchanged. This research program focusses on the detailed ;investigation of membrane-protein as well as phospholipid-protein interactions, using a combinationof chemical, spectroscopic, and kinetic techniques. Selective chemical modification is used to identify amino acyl residues in the phospholipid binding site and the membrane attachment domain of the transfer protein. Fluorescence spectroscopy and the associated quenching, polarization, and lifetime of chromophores is used to define the physicochemical nature of membrane-protein and phospholipid-protein complexes. Circular dichroism is used in an analogous manner, with the further advantage of stopped-flow analysis. The magnitude and transbilayer distribution of various phospholipid pools, particularly phsophatidylinositol, in artificial single bilayer vesicles of different size and composition are determined and correlated with the accessibility to phospholipid transfer proteins. Lateral phase separations are generated in model membranes and explored as possible sites of altered protein-catalyzed phsopholipid transfer activity. The results should help describe the mechanism of action of phsopholipid transfer proteins and to define the biological roles of these proteins in phosphatidylinositol metabolism and membrane assembly and homeostasis.
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