CALMODULIN ACTIVATION AND INHIBITION OF SKELETAL-MUSCLE CA2+ RELEASE CHANNEL (RYANODINE RECEPTOR)

The calmodulin-binding properties of the rabbit skeletal muscle Ca2+ r elease channel (ryanodine receptor) and the channel's regulation by ca lmodulin were determined at less than or equal to 0.1 PM and micromola r to millimolar Ca2+ concentrations. [I-125]Calmodulin and [H-3]ryanod ine binding to sarcoplasmic reticulum (SR) vesicles and purified Ca2release channel preparations indicated that the large (2200 kDa) Ca2release channel complex binds with high affinity (K-D = 5-25 nM) 16 ca lmodulins at less than or equal to 0.1 mu M Ca2+ and 4 calmodulins at 100 mu M Ca2+. Calmodulin-binding affinity to the channel showed a bro ad maximum at pH 6.8 and was highest at 0.15 M KCl at both less than o r equal to 0.1 mu M and 100 mu M Ca2+. Under conditions closely relate d to those during muscle contraction and relaxation, the half-times of calmodulin dissociation and binding were 50 +/- 20 s and 30 +/- 10 mi n, respectively. SR vesicle-Ca-45(2+) flux, single-channel, and [H-3]r yanodine binding measurements showed that, at less than or equal to 0. 2 mu M Ca2+, calmodulin activated the Ca2+ release channel severalfold . At micromolar to millimolar Ca2+ concentrations, calmodulin inhibite d the Ca2+-activated channel severalfold. Hill coefficients of similar to 1.3 suggested no or only weak cooperative activation and inhibitio n of Ca2+ release channel activity by calmodulin. These results sugges t a role for calmodulin in modulating SR Ca2+ release in skeletal musc le at both resting and elevated Ca2+ concentrations.