Muscle contraction is a fundamental physiological process. Two types of calcium channels, the voltage-gated Ca2+ (Cav) channels on the plasma membrane and the huge high-conductance Ca2+ channel RyR on the ER membrane of myocytes, are responsible for the initiation of muscle contraction through a mechanism known as 'excitation-contraction (EC) coupling'. In skeletal muscle, the conformational changes of Cav1.1 upon membrane depolarization are coupled to the activation of the downstream RyR1 through physical associations. In cardiocytes, RyR2 is activated by the Ca2+ influx mediated by Cav1.2, a mechanism known as 'calcium-induced calcium release'. The rapid release of Ca2+ ions from SR through RyR1/2 triggers a cascade of events that lead to muscle contraction.The goal of this proposal is to achieve an improved structural and mechanistic understanding of the EC coupling in cardiac muscle, which is essential for the function of heart. My project is focused on the two important proteins during this process, Cav1.2 and RyR2. My first specific aim is to study the structural basis for the modulation of Cav1.2 channels by representative drugs. My second specific aim is structural investigation of the luminal calcium sensing mechanism of RyR2. My third specific aim is structural studies of the pharmacology of RyR2. To achieve these goals, the first step is to obtain decent protein sample. I plan to try recombinant expression in mammalian cell expression system, and then optimize the purification strategy, including the detergent, affinity tag buffer condition and so on. After getting the optimal protein sample, I will try to solve the structure by single particle cryo EM method.The objectives to be achieved is structures of Cav1.2 and drugs, which will reveal the molecular basis for the specificity and potency of various drugs, can serve as templates to facilitate ligand optimization and future drug discovery. The objectives for RyR2 are the high resolution structure with the luminal calcium sensor and also the complex structure of RyR2 and drugs. it is of great clinical significance to study the luminal calcium sensing mechanism of RyR2. The results will also provide the structure basis for the pharmacology and shed light on the advanced accurate drug development target on RyR2.Taking together, they will provide an improved structural and mechanistic understanding of the EC coupling in cardiac muscle, which is essential for the function of heart.
|Program type||Postdoctoral Fellowship|
|Effective start/end date||01/01/2019 → 09/07/2019|