Elsevier

Cell Calcium

Volume 15, Issue 4, April 1994, Pages 305-316
Cell Calcium

Calcium-dependent block of ryanodine receptor channel of swine skeletal muscle by direct binding of calmodulin

https://doi.org/10.1016/0143-4160(94)90070-1Get rights and content

Abstract

The interaction of the Ca2+ binding protein calmodulin (CaM) with the ryanodine receptor of the sarcoplasmic reticulum (SR) of pig skeletal muscle was investigated by [3H]-ryanodine binding, planar bilayer recordings, and rapid filtration of 45Ca2+-loaded SR. Inhibition of [3H]-ryanodine binding by CaM was phosphorylation-independent, had an IC50 of approximately 0.1 μM and was optimal at 10 μM Ca2+. CaM also inhibited [3H]-ryanodine binding to CHAPS-solubilized and purified ryanodine receptors, suggesting a direct CaM-ryanodine receptor interaction. In single channel recordings, CaM blocked Ca2+ release channels in a Ca2+-dependent manner by decreasing the number of open events per unit time without affecting the mean open time or unitary channel conductance. Rapid filtration of 45Ca2+ passively loaded into SR vesicles showed that CaM blocked Ca2+ release within milliseconds of exposure of SR to a Ca2+ release medium containing 10 μM CaM. In controls, an increase in extravesicular Ca2+ from 7 nM to 10 μM resulted in a release of 47 ± 10% of the 45Ca2+ in 20 ms. CaM reduced the release to 23 ± 12% in the same period. These results are compatible with a direct mechanism of Ca2+ release channel blockade by CaM and suggest that CaM could play a significant role in the inactivation of SR Ca2+ release during excitation-contraction coupling.

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    On leave from the Laboratorio de Fisiología, Universidad del Bío-Bío, Chillán, Chile.

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