o-phthalaldehyde activates the Ca2+ release mechanism from skeletal musclesarcoplasmic reticulum

omicron -Phthalaldehyde (OPA) is a bifunctional reagent that forms an isoin dole derivative by reacting with cysteine and lysine residues separated by approximately 0.3 nm. OPA inhibits sarcoplasmic reticulum (SR) Ca2+-ATPase activity at low micromolar concentrations and induces Ca2+ release from act ively loaded SR vesicles by activating the ryanodine receptor from fast twi tch skeletal muscle. Both ryanodine binding and single-channel activity sho w a biphasic concentration dependence, At low OPA concentrations (< 100 muM ), ryanodine binding and single channel activity are stimulated, while at h igher concentrations, a time-dependent sequential activation and inhibition of receptor binding is observed. Activation is characterized by a Ca2+-ind ependent increase in maximal receptor occupancy, Data are presented to supp ort a model in which Ca2+ channel and ryanodine binding activity are enhanc ed due to an intramolecular cross-linking of nearby lysine and nonhyperreac tive cysteine residues. OPA complexation with endogenous lysine residue(s) is critical for receptor activation. (C) 2001 Academic Press.