Calmodulin is a widely distributed, multifunctional calcium binding protein that plays a role in many diverse biological events, including neurotransmission, the development and structure of cells, cellular metabolism and various secretary processes. The overall objective of this research program is to inquire further into the role calmodulin plays in biological processes with the ultimate goal of selectively altering its activity. The strategy is to develop and study pharmacological probes that interfere with discrete actions of calmodulin. This problem will be addressed from different approaches: (a) To study certain peptide and non-peptide inhibitors of calmodulin with the aim of not only uncovering more potent and selective inhibitors, but also to identify calmodulin inhibitors that act at different sites on the calmodulin molecule; and (b) to develop agents that act not on calmodulin itself, but rather on the calmodulin binding sites of specific enzymes that are activated by calmodulin. The types of compounds that will be studied are irreversibly bound phenothiazine-calmodulin complexes, peptide-calmodulin complexes and fragments of calmodulin or other calcium binding proteins. The relative selectivity of these various inhibitors of calmodulin will be assessed by determining their effects on certain calmodulin-sensitive enzymes and calmodulin-dependent processes. The enzymes that will be studied initially will be the calmodulin-sensitive forms of phosphodiesterase, adenylate cyclase, ATPase and protein kinase. To determine further the drug selectivity, comparisons will be made with calmodulin-insensitive forms of the enzymes and with phospholipid-sensitive forms of the enzymes. To study the functional correlates of these biochemical studies and to determine whether we can discern which specific calmodulin-dependent enzyme is involved in a specific biological event, we will measure the effects of the inhibitors on certain biological process, including: (a) their effects on the contractility of intestinal smooth muscle, a study that may provide new insights into the regulation of gastrointestinal motility and blood pressure; and (b) their effects on the development and maturation of oocytes, processes that may have implications not only in embryogenesis, but also in other general aspects of cellular growth and differentiation.
