Relationship of low affinity [3]ryanodine binding sites to high affinity sites on the skeletal muscle Ca2+ release channel.
Both high and low affinity binding sites for [3H]ryanodine exist in sarcoplasmic reticulum membranes derived from rabbit skeletal muscle. Negatively cooperative binding of [3H]ryanodine at one of four initially identical sites cannot account for some of the kinetic features of the binding to high and low affinity sites. The presence of excess unlabeled ryanodine greatly slows the rate at which [3H]ryanodine bound at the high affinity site dissociates. An examination of the rate of dissociation of [3H]ryanodine bound at increasing [3H]ryanodine concentrations reveals the existence of a second site, occupied only at high ligand concentrations. The occupation of this site correlates well with the conversion of the high affinity site from a site with a dissociation rate constant of approximately 0.0025 min-1 to one with a dissociation rate constant of less than 0.00025 min-1. The low affinity site itself has a dissociation rate constant of 0.013 min-1 and dissociation from this site is unaffected by the presence of 100 microM unlabeled ryanodine. These data suggest that the two binding sites are different but are either allosterically or sterically coupled. Association experiments support this interpretation. Low affinity binding sites for [3H]ryanodine exist in transverse tubule (t-tubule) as well as sarcoplasmic reticulum membranes. High concentrations of both ryanodine and ruthenium red inhibit the binding of [3H]PN200-110 to the dihydropyridine-binding protein in t-tubule membranes. Whether the low affinity site in t-tubule membranes is related to that found in sarcoplasmic reticulum membranes is not yet known.
Wang
J P
JP
Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, Texas 77030.
Needleman
D H
DH
Hamilton
S L
SL
D000818:Animals; D001665:Binding Sites; D015220:Calcium Channels / Q000378:metabolism*; D007700:Kinetics; D009132:Muscles / Q000378:metabolism*; D011817:Rabbits; D012433:Ryanodine / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D014316:Tritium
D015220:Calcium Channels; D014316:Tritium; D012433:Ryanodine