调控剂的电子传递性质对骨骼肌型钙释放通道的调控效应研究

肌质网(sarcoplasmic reticulum，SR)中的钙释放通道利阿诺定受体(ryanodine receptor，RyR)是调控胞浆钙离子浓度的重要蛋白，其活性受多种调控剂影响．调控剂的不同电子传递性质可能作用于RyR的功能性巯基，进而影响其门控状态．了解具有不同电子传递性质的调控剂影响钙通道的作用机制具有重要意义．本研究采用光子相关光谱法(PCS)、CPM(7-二乙 基-3-(4′-马来酰亚胺苯基)4-甲基香豆素)荧光标记法及[³H]-ryanodine结合等实验，分别检测多种调控剂对RyR1的蛋白及复合体粒度分布、自由巯基量及对通道状态的影响，利用光漂白法检测各调控剂的电子传递性质．结果显示，激活剂和巯基氧化剂具有类似电子受体的性质并产生相似作用，即自由蛋白粒度增加，自由巯基量减少，具有激活通道作用；抑制剂和巯基还原剂则具有类似电子供体的性质，作用效果相反．

The Effects of Ryanodine Receptor Modulators on The Calcium Release Channel Functions According to Their Electron Transfer Properties

Ryanodine receptor (RyR) locating in the sarcoplasmic reticulum(SR) is one of the most important calcium release channels. It is known that RyR is rich in functional thiol groups, which play crucial roles in controlling channel gating, therefore the protein is quite sensitive to the redox environment. The main purpose of this research is to understand how the RyR1 modulators in general affect the channel gating according to their electron transfer properties. In this research, methods such as CPM labeling, photon correlation spectroscopy (PCS) and [³H]-ryanodine binding experiments were used to detect the change of free thiols' population, the diameter and distribution of protein complex and the gating state of RyR1. Furthermore, we used photon bleaching assay to determine the electron transfer property of the modulators. The results show that the channel activators and thiol oxidants appear to be electron acceptors that affect RyR1 in the same manner, which is to decrease the free thiols on RyR1 and to increase the size of the protein crosslinking clusters. In contrary, channel inhibitors and thiol reductants appear to be electron donors and their effects on RyR1 are opposite to activators and oxidants. Therefore, we propose that the similar electron transfer properties of channel modulators results in the similar effects on RyR1 by changing the local redox environment around the active thiols, inducing changes in thiol/disulfide balance and RyR1 gating.