9317684 Welch Ryanodine is a plant alkaloid which binds to a specific receptor,an intracellular calcium release channel, first identified at the triad junction of skeletal muscle. Subsequently, ryanodine receptors (RyR) have been found in many non-muscle cells including cells of the central nervous system. Ryanodine has been shown to be a powerful effector of calcium regulation in muscle. Although the ryanodine molecule is complex, important stearic and electrostatic interactions can be localized to relatively small, specific regions of the molecule. The overall goal of this project is to elucidate the relationship between ligand structure and the biological activities of the ryanoids. The aims are to (1) develop biologically active ryanoids with minimal structure, (2) develop agents which effect specific alterations in RyR properties, (3)define the mammalian skeletal muscle ryanodine binding site, (4)lay the groundwork for development of RyR isoform-specific agents and (5) define endogenous effectors. This project will focus on rabbit skeletal muscle RyR. Approaches will be: (I) New ryanodine analogs will be developed. (II) The biological activity of these compounds will be assessed by measurements of ligand binding to the RyR, the modulation of ion channel activity of the RyR using ionic flux and planar lipid bilayer single channel measurements, and functional effects on intact cells. (III) The conformations of the ryanoids will be deduced from the covalent structures by molecular mechanics, molecular dynamics and quantum mechanics. The deduced conformations will be compared to NMR or crystallographic data whenever possible (for example, the X-ray and NMR structures of ryanodine have been solved). A comparative molecular field analysis (ComFA) will be produced for selected analogs, and correlations between the stearic and electrostatic fields computed to discern the relationship between the biological activities and the structure of the ryanodine a nalogs. (IV) The results of the CoMFA will be used to design new ryanoids for testing, refinement and correction of hypotheses. %%% Calcium ion regulates many critical cellular functions such as muscle contraction. The levels of calcium free in cells are precisely regulated by physiological control mechanisms. Ryanodine is a plant alkaloid which has a powerful effect on calcium regulation in cells and can serve as a useful tool in studying calcium control mechanisms. Ryanodine acts by binding to a specific receptor, an intracellular calcium release channel. Ryanodine receptors were first identified in skeletal muscle, but have also been found in many non-muscle cells including cells of the central nervous system. Although the ryanodine molecule is complex, its functional interactions with its receptor can be localized to relatively small, specific regions of the molecule. The overall goal of this project is to elucidate the relationship between the structure and the biological activities of the ryanoids. This will result in the development of important new probes which will enhance our understanding of cellular calcium regulation. ***
