STRUCTURAL DETERMINANTS OF HIGH-AFFINITY BINDING OF RYANOIDS TO THE VERTEBRATE SKELETAL-MUSCLE RYANODINE RECEPTOR - A COMPARATIVE MOLECULAR-FIELD ANALYSIS

Ryanodine binds to specific membrane proteins, altering the calcium pe rmeability of intracellular membranes. In this study 19 ryanoids were isolated or synthesized and the structures correlated to the strength of binding to vertebrate skeletal muscle ryanodine receptors. Global m inima were determined by employment of molecular mechanics and dynamic s augmented by systematic searching of conformational space. Overall, steric and electrostatic factors contribute about equally to the diffe rences in the experimentally determined dissociation constants. The do minant electrostatic interaction is localized to a hydroxyl group in a n apolar region of the molecule. The pyrrole and isopropyl groups loca ted together at one pole of the molecule have the greatest effect on s teric interactions between ligand and receptor. We suggest ryanodine b inds to the receptor with the pyrrole and isopropyl groups buried deep inside a cleft in the protein. This arrangement places special import ance on the conformation of the pyrrole and isopropyl groups. In contr ast, the opposite pole appears to be positioned at the entrance of the binding pocket because bulky adducts placed in the 9 position of ryan odine alter binding minimally. For example, a fluorescent ryanodine ad duct was synthesized which has a dissociation constant close to that o f ryanodine. Detailed examination reveals subtle interactions between ryanoid and receptor. In many cases, the major factors altering the st rength of binding were found to be conformational alterations in the m olecule remote from the site of covalent modification.