[3H]ryanodine as a probe of changes in the functional state of the Ca(2+)-release channel in malignant hyperthermia.
The defect in malignant hyperthermia (MH) alters the binding of [3H]ryanodine to the Ca(2+)-release channel by increasing its apparent affinity for the binding site. In sarcoplasmic reticulum (SR) membranes from both normal and mutant pigs the apparent Kd is dependent on a number of parameters. Adenosine 5'-(beta,gamma-methylene)triphosphate, ionic strength, and Ca2+ each increase the apparent affinity of the binding site for [3H]ryanodine. Equilibrium and kinetic evaluation of the binding of [3H]ryanodine to these membranes demonstrates that the MH defect in pigs increases the apparent affinity of the membranes for [3H]ryanodine by increasing the amount of high affinity relative to low affinity binding sites. Both the association and dissociation of [3H]ryanodine with all three types of membranes (normal, heterozygous MH, homozygous MH) are characterized by two or more components, with the relative ratios of these components altered by the MH defect. These findings suggest that the observed Kd is the weighted average of the binding of ryanodine to two or more interconvertible states of the channel. Dilution of [3H]ryanodine bound to normal membranes at high Ca2+ into low Ca2+ solutions enhances the rate of dissociation. This conversion occurs to a much lesser extent with MH membranes, suggesting that the MH defect may alter the rate at which the high affinity form of the protein converts to the low affinity form.
Hawkes
M J
MJ
Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, Texas.
Nelson
T E
TE
Hamilton
S L
SL
D000818:Animals; D001706:Biopsy; D002110:Caffeine / Q000494:pharmacology; D015220:Calcium Channels / Q000502:physiology*; D006579:Heterozygote; D066298:In Vitro Techniques; D007700:Kinetics; D008305:Malignant Hyperthermia / Q000235:genetics / Q000378:metabolism*; D009119:Muscle Contraction; D009132:Muscles / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology; D012433:Ryanodine / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D013552:Swine; D014316:Tritium
D015220:Calcium Channels; D014316:Tritium; D012433:Ryanodine; D002110:Caffeine