Nicotinic receptors (nAChR) play a key role in synaptic transmission at the neuromuscular junction. Several mutations in the human muscle nAChR subunits, which disrupt the mechanism of activation, cause rnyasthenia gravis syndrome. The nature of conformational changes induced by agonist, which mediate the activation in nAChRs, remains unresolved. In the present project, I will utilize fluorescence spectroscopy to monitor the conformational dynamics within nAChRs in real time. The overall goal of the present proposal is to gain expertise in spectroscopic mapping of the conformational dynamics induced by agonist in the muscle nAChR.
The specific aims are 1) to characterize the nature of conformational changes of TM2 domain detected by the MTSR fluorophore at the 19' position, 2) to test the hypothesis that 19' residue may contribute to the gating process in a subunit-dependent manner, 3) to explore the molecular motion in the TM2 domain of various subunits by FRET, and finally 4) to monitor conformational changes in both N-terminus loops 2 and 7 and the TM2-TM3 loop. The proposed spectroscopic studies will help us to understand the mechanism of activation in nAChRs and design better therapeutics. ? ?
