SYNTHETIC INHIBITORS OF MEMBRANE ANCHOR PHOSPHOLIPASES

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In the past four years increasing attention has been focused on a distinct type of membrane anchors of cell surface proteins, the glycosyl phosphatidylinositols (GPI). Among more than 40 of GPI-anchored proteins identified to date are various cell surface antigens, tumor markers, receptor systems, complement regulatory factors, cell adhesion molecules and membrane enzymes. The biosynthesis, metabolism and biological functions of glycolipid anchors are under active investigation. The release of surface proteins and GPI components with putative signal transduction properties by GPI-specific phospholipase C (GPI-PLC) and D (GPI-PLD) have been described in several parasitic and mammalian cell systems. Thus, there is a timely need for specific inhibitors and biochemical probes of these two phospholipases to facilitate the biological study of many important GPI-anchored proteins. Our preliminary data have shown that a unique GPI component previously synthesized in our laboratory selectively inhibit the trypanosome GPI- PLC, but not a PI-PLC, in a cell-free system at 1 mM in a stereospecific manner. We propose to use derivatives of this component as core structures to maintain GPI-specificity and develop more potent inhibitors of both GPI-PLC and a mammalian GPI-PLD. Various substituted, noncleavable and transition state analogs of the respective substrates will be synthesized. The optimal conformation of active inhibitors will be analyzed by NMR, X-ray and computer modeling. Related photoactivable biochemical probes will be prepared for a collaborator to characterize the active site of these enzymes. The effect of specific inhibitors on (1) the biosynthesis and metabolism of GPI-anchored proteins in Trypanosome brucei and Entamoeba histolytica, and (2) the signal transduction process of interleukins in human lymphocytes will also be investigated with collaborators towards the development of potential antiparasitic agents and novel immunomodulators.