This award will support collaborative research in biochemistry between Dr. Daniel Watterson, Vanderbilt University School of Medicine, and Dr. Jacques Haiech, Centre de Recherches de Biochimie Macromoleculaire, Montpellier, France. The objective of this research is to study the molecular basis of calcium and cylic nucleotide signal transduction. Calcium and cyclic AMP are two important second messengers in eukaryotic cells. Variations of their concentrations serve as intracellular signals. Appropriate receptors (calcium and cAMP binding proteins) are key elements in the detection and management of these signals. Among the calcium binding proteins involved in the calcium signal management of eukaryotic cells, calmodulin appears to be a prototypical calcium sensor. Upon calcium binding, this protein modifies its conformation and interaction with target proteins, thus transmitting the information to another element in a molecular cascade that results in a cellular event (e.g., contraction, exocytosis). Among the target proteins is a calmodulin sensitive adenylate cyclase. The objectives of the proposed research projects are: 1) To study the mechanisms of calcium binding to calmodulin by using a site-directed mutagenesis and protein engineering approach. The investigators will design isofunctional mutants with a single tryptophan that will be used as a reporter group in fluorescence studies; 2) To isolate a cDNA coding for the adenylate cyclase from Chlamydomonas. This cDNA _____________ will be used to produce an engineered protein in order to study the interaction between the adenylate cyclase and calmodulin. Both the U.S. and French investigators have significant expertise in the proposed research area. Dr. Watterson's laboratory has developed methods for the mutagenesis of calmodulin, while Dr. Haiech's laboratory has the facilities for growing the expressed proteins in quantity and for the physical analysis of the resulting calmodulins.
