Interactions of a reversible ryanoid (21-amino-9alpha-hydroxy-ryanodine) with single sheep cardiac ryanodine receptor channels.
The binding of ryanodine to a high affinity site on the sarcoplasmic reticulum Ca2+-release channel results in a dramatic alteration in both gating and ion handling; the channel enters a high open probability, reduced-conductance state. Once bound, ryanodine does not dissociate from its site within the time frame of a single channel experiment. In this report, we describe the interactions of a synthetic ryanoid, 21-amino-9alpha-hydroxy-ryanodine, with the high affinity ryanodine binding site on the sheep cardiac sarcoplasmic reticulum Ca2+-release channel. The interaction of 21-amino-9alpha-hydroxy-ryanodine with the channel induces the occurrence of a characteristic high open probability, reduced-conductance state; however, in contrast to ryanodine, the interaction of this ryanoid with the channel is reversible under steady state conditions, with dwell times in the modified state lasting seconds. By monitoring the reversible interaction of this ryanoid with single channels under voltage clamp conditions, we have established a number of novel features of the ryanoid binding reaction. (a) Modification of channel function occurs when a single molecule of ryanoid binds to the channel protein. (b) The ryanoid has access to its binding site only from the cytosolic side of the channel and the site is available only when the channel is open. (c) The interaction of 21-amino-9alpha-hydroxy-ryanodine with its binding site is influenced strongly by transmembrane voltage. We suggest that this voltage dependence is derived from a voltage-driven conformational alteration of the channel protein that changes the affinity of the binding site, rather than the translocation of the ryanoid into the voltage drop across the channel.
Tanna
B
B
Cardiac Medicine, National Heart & Lung Institute, Imperial College of Science, Technology & Medicine, London SW3 6LY, United Kingdom.
Welch
W
W
Ruest
L
L
Sutko
J L
JL
Williams
A J
AJ
D000818:Animals; D003600:Cytosol / Q000737:chemistry / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004553:Electric Conductivity; D004594:Electrophysiology; D015640:Ion Channel Gating / Q000502:physiology; D007700:Kinetics; D008564:Membrane Potentials / Q000502:physiology; D009206:Myocardium / Q000737:chemistry*; D012433:Ryanodine / Q000031:analogs & derivatives* / Q000494:pharmacology; D019837:Ryanodine Receptor Calcium Release Channel / Q000502:physiology*; D012519:Sarcoplasmic Reticulum / Q000737:chemistry / Q000378:metabolism; D012756:Sheep
C113821:21-amino-9-hydroxyryanodine; D019837:Ryanodine Receptor Calcium Release Channel; D012433:Ryanodine