The nicotinic acetylcholine receptor channel (AChR) is an allosteric membrane protein that mediates neuromuscular transmission in vertebrates. Experiments are proposed regarding the kinetic properties of wild type and mutant mouse recombinant AChR, probed using single-channel electrophysiology. Three areas that are of fundamental importance with regard to understanding synaptic receptor channels are addressed: agonist binding, channel gating, and desensitization. One goal of this proposal is to understand the mechanisms of molecular recognition at the transmitter binding sites. Experiments are proposed to study the nature of the agonist-protein interactions with both high and low affinity conformations of the AChR. The second objective is to use linear free energy relationships to obtain a snapshot of the gating transition state. These experiments will report the order of events during the closed-open allosteric transition.
The third aim i s to understand the molecular basis of desensitization. Experiments are proposed to make high-resolution measurements of desensitization recovery rate constants in both wild type and mutant AChR. The results of the experiments will provide a deep, mechanistic understanding of AChR both at normal synapses, and at synapses that have been compromised by neuromuscular disease.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Method to Extend Research in Time (MERIT) Award (R37)
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Special Emphasis Panel (ZRG1-MDCN-3 (01))
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Silberberg, Shai D
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State University of New York at Buffalo
Schools of Medicine
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Purohit, Prasad; Auerbach, Anthony (2013) Loop C and the mechanism of acetylcholine receptor-channel gating. J Gen Physiol 141:467-78
Jadey, Snehal; Purohit, Prasad; Auerbach, Anthony (2013) Action of nicotine and analogs on acetylcholine receptors having mutations of transmitter-binding site residue ýýG153. J Gen Physiol 141:95-104
Nayak, Tapan K; Auerbach, Anthony (2013) Asymmetric transmitter binding sites of fetal muscle acetylcholine receptors shape their synaptic response. Proc Natl Acad Sci U S A 110:13654-9
Gupta, Shaweta; Purohit, Prasad; Auerbach, Anthony (2013) Function of interfacial prolines at the transmitter-binding sites of the neuromuscular acetylcholine receptor. J Biol Chem 288:12667-79

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