Ondrias K, Brillantes A M, Scott A, Ehrlich B E, Marks A R
Department of Medicine, Mount Sinai School of Medicine, New York 10029, USA.
Soc Gen Physiol Ser. 1996;51:29-45.
The calcium-release channel/ryanodine receptor of the sarcoplasmic reticulum is a 2.3 million-D structure required for intracellular calcium release during excitation-contraction coupling in skeletal muscle. This structure is the largest ion channel characterized to date and is composed of four 565,000-D ryanodine receptors plus four molecules of FKBP12. In the present study we describe the single channel properties of the cloned expressed ryanodine receptor, with and without FKBP12, reconstituted into planar lipid bilayers with Ca as the charge carrier. The conductance for Ca (luminal, 53 mM/cytoplasmic, 10 microM) was 103 pS for the cloned expressed RyR and for the native channel from rabbit skeletal muscle. Conductance through the channel was Ca dependent: A decrease in the Ca gradient to luminal 10.6/cytoplasmic 10 microM reduced conductance to 68 pS for both the cloned and native RyR. The recombinant ryanodine receptor consistently behaved like the native skeletal muscle channel in terms of activation by caffeine, calcium, and ATP; inhibition by ruthenium red; and modulation by ryanodine. In the absence of FKBP12, the cloned expressed RyR exhibited multiple subconductance states and addition of FKBP12 reduced the frequency of subconductance states. These results show that with Ca as the charge carrier, the single channel properties of the cloned expressed RyR plus FKBP12 are essentially the same as those of the native channel.
肌浆网的钙释放通道/雷诺丁受体是一种230万道尔顿的结构，在骨骼肌兴奋-收缩偶联过程中，细胞内钙释放需要该结构。这种结构是迄今为止已鉴定的最大离子通道，由四个56.5万道尔顿的雷诺丁受体加上四个FKBP12分子组成。在本研究中，我们描述了克隆表达的雷诺丁受体（有或没有FKBP12）的单通道特性，该受体被重组到以Ca作为电荷载体的平面脂质双分子层中。对于克隆表达的RyR和来自兔骨骼肌的天然通道，Ca（管腔，53 mM/细胞质，10 microM）的电导为103 pS。通过该通道的电导依赖于Ca：将Ca梯度降低至管腔10.6/细胞质10 microM时，克隆的和天然的RyR的电导均降低至68 pS。重组雷诺丁受体在咖啡因、钙和ATP激活、钌红抑制以及雷诺丁调节方面，始终表现得与天然骨骼肌通道一样。在没有FKBP12的情况下，克隆表达的RyR表现出多个亚电导状态，添加FKBP12可降低亚电导状态的频率。这些结果表明，以Ca作为电荷载体时，克隆表达的RyR加FKBP12的单通道特性与天然通道基本相同。
