Structural Determinants of the Biological Activity of Ryanodine

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The ryanodine receptor, situated in the triad junction of muscle cells, normally functions as the calcium-release channel that couples depolarization of the sarcolemma with calcium release from the sarcoplasmic reticulum to trigger sarcomere contraction. The receptor is also found in several other cell types where its function in not as clear. The receptor is so-named because of its ability to bind the plant alkaloid, ryanodine, which stabilizes the open channel. The wide distribution of the receptor in muscle and other tissues and the complexity by which channel activity is modulated by a variety of agents strongly suggest a fundamental role for the receptor in cellular function. At present it is not known (a)what the molecular mechanisms are that control channel activity of the receptor, (b) the physiological significance of the receptor in non-muscle cells, and (c) whether there are endogenous ligands that bind to the ryanodine site and affect the channel. The long-term goal of this project is to understand the molecular mechanism of ryanodine modulation of channel activity and the role of the receptor in the intact cell. The basic strategy is to combine directed chemical synthesis of ryanodine analogs, the testing of these analogs for their biological activities, and the computer-assisted modeling of the molecular structure of the receptor based on these activities. The specific aims of the project are the following:1. To synthesize and test ryanodine analogs to determine the minimal structural factors that define their agonist and antagonist properties.2. To map the known high and low affinity ryanodine binding sites and to test their functional properties.3. To map those loci of the binding sites that induce the open and closed states of the channel.The project is a continuing collaborative effort of four laboratories, which will contribute expertise in organic synthesis, membrane electrophysiology, pharmacology and computer modeling.