There is a tremendous current and growing interest in the role of membrane phosphoinositides in cell biology, especially in relation to the transmission of Ca2+ mediated signals which are generated when cell-surface receptors are activated by hormones, neurotransmitters, growth factors, and other agonists, in relation to the mechanism of malignant transformations induced by oncogenes, and, the activation of kinase C. Samples of pure phosphoinositides and analogs are important for research and as potential therapeutic agents. Practical methods for their preparation by synthesis are not available. The overall objective of this proposal is to develop a general, simple and practical method for the synthesis of distinct molecular species of phosphatidylinositol (PI), phosphatidylinositol-4-phosphate (PIP), and, phosphatidylinositol-4,5-bisphosphate (PIP2), each carrying fattyacyl residues of choice in positions 1', 2'. The strategy is to replace the mixtures of fattyacyls in natural PI and derivatives with any desired fattyacyls by a deacylation - reacylation cycle, or, by removal and reintroduction of the complete diglyceride residue. The synthesis depends critically on an innovative application of lipases in a composite chemical and enzymatic sequence. During Phase I, this critical aspect, and the pre-requisite protection and deblocking of the inositol hydroxyls will be evaluated for the preparation of semi-synthetic 1',2'-diacyl-sn-glycerophosphoinositols from PI isolated from soylecithin, so as to determine the feasibility of the general strategy.
