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Positive cooperativity of ryanodine binding to the calcium release channel of sarcoplasmic reticulum from heart and skeletal muscle



Positive cooperativity of ryanodine binding to the calcium release channel of sarcoplasmic reticulum from heart and skeletal muscle



Biochemistry 28(4): 1686-1691



Ryanodine is a specific ligand for the calcium release channel which mediates calcium release in excitation .sbd. contraction coupling in muscle. In this study, ryanodine binding in sarcoplasmic reticulum from heart muscle and skeletal muscle is further compared and correlated with function. The new findings include the following: (1) Two types of binding, high affinity (KD1 .apprx. 5.sbd.10 nM) and low affinity (Kd2 .apprx. 3 .mu.M), can now be discerned for the skeletal muscle receptor. KD1 is approximately the same as and KD2 of similar magnitude to that previously reported for heart. (2) The dissociation rates for the high-affinity binding have been directly measured for both heart and skeletal muscle (t1/2 .apprx. 30.sbd.40 min). These rates are more rapid than previously reported (t1/2 .apprx. 14 h). (3) KD1's obtained from the ratio of the dissociation and association rate constants agree with the dissociation constant measured by equilibrium binding Scatchard analysis. (4) Ryanodine binding to the low-affinity site can be correlated with a decrease in the dissociation rate constant (k-1) of the high-affinity site, and thereby in the apparent dissociation constant (KD1). The inhibition constant (KI) for inhibiting the high-affinity off rate obtained from a double-reciprocal plot of the change in off rate vs [ryanodine] is practically the same in heart (0.66 .mu.M) and skeletal muscle (0.64 .mu.M) and in the range of the KD2. The binding of cold ryanodine to the low-affnity site appears to lock the bound [3H]ryanodine onto the high-affinity site rather than to exchange with it. Thus, in this sense, the ryanodine receptor exhibits "positive cooperativity". For both heart and skeletal muscle, the binding of ryanodine to the high-affinity site has previously been correlated with the pharmacologic effect of locking of the channels in the "open" state. We now find that binding to the low-affinity site in skeletal muscle can be correlated with the pharmacologic effect of closing the channel, comparable to that which we have previously reported for heart. The Km for closing of the channel in heart (1.1 .mu.M) and that in skeletal muscle (3.7 .mu.M) are in the same range as the KI and KD2. These studies indicate that the low-affinity binding site mediates the closing of the channel and is responsible for the positive cooperativity. The binding characteristics of ryanodine to the receptor (calcium release channel) and their functional correlates (Km and KI) are similar in heart and skeletal muscle.

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Accession: 007669144

Download citation: RISBibTeXText

PMID: 2541762

DOI: 10.1021/bi00430a039


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