The Electron Transfer Properties Effects Of Ryanodine Receptor Modulators On The Channel Proteins

Ryanodine receptor (RyR) which located in sarcoplasmic reticulum (SR) is one of the most important calcium release channels. The activity of RyR can be modulated by many effectors, including redox reagents and non-redox modulators such as tetracaine, caffeine and its congeners. It's known that RyR is rich in functional thiol groups, which play crucial roles in controlling channel gating. Therefore it's quite sensitive to the redox environment. Previous studies have shown that thiol oxidants have similar effects on RyRl, such as stimulating the Ca2+release, reducing the number of free thiols on RyR1 and so on. On the contrary, most of the thiol reductants behave in the opposite manner. It's also found that modulators with weak redox properties could also affect the number of free thiols on RyR1. Here come to the main purpose of this research, that is to find out how the modulators affect the channel proteins by their electron transfer properties.In this research, methods such as CPM labeling, surface enhanced Raman spectrum, photon correlation spectroscopy (PCS) and [3H]-Ryanodine binding experiments to dectect the change of free thiols on RyR1, the signals of thiols and disulfide, the diameter and distribution of proteins and the gating state of RyRl. Furthermore, we use photon bleaching assay to measure the electron transfer properties of the modulators and then compare with the result of quantum calculations.The results show that the channel activators and thiol oxidants appears to be electron accpetors, they can affect RyR1 in the same manner, that is to decrease the free thiols on RyR1, increase the signal of disulfides and the distribution of the clusters. In contrary, channel inhibitors and thiol reductants appears to be electron donors and their effects on RyR1 are opposite to activators and oxidants. Now we can conclude that the electron transfer properties of channel modulators can affect RyRl by shifting the local redox environment of the hyperreactive thiols, inducing changes in thiol/disulfide balance and then result in the changes of RyR1 gating.