Dynamics Of Ligand Gated Ion Channels

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PROJECT SUMMARY/ABSTRACT Glutamate receptors mediate the spread of excitation in the mammalian central nervous system, and ultimately control physiological functions such as movement and cognition. They are classified into four subfamilies: N-methyl-D-aspartate (NMDA), -amino-5-methyl-3-hydroxy-4-isoxazole propionate (AMPA), kainate, and delta receptors. All four subtypes are topologically similar, comprising four protein subunits held together as a dimer-of-dimers. However, their gating characteristics and mechanisms are unique, reflecting distinct roles in synaptic transmission. The goal of my laboratory is to understand the mechanisms that underlie the functional fine-tuning of each of these subtypes. Using a multipronged approach that combines biochemical, electrophysiological, fluorescence, and computational methods, we have already identified conformational transitions that are critical for mediating activation and inhibition of AMPA and NMDA glutamate receptors. However, a direct understanding of how such structural changes in one part of the protein control changes in other parts, and how they dictate functional properties, remains unknown. We propose to develop and use multicolor single-molecule fluorescence resonance energy transfer (FRET) to draw structure–function correlations between different segments of each glutamate receptor subtype. We will also focus our efforts on understanding the less well-studied kainate and delta subtypes. Specifically, we will investigate the mechanisms underlying activation and modulation of an abundant form of the kainate receptor by agonists, ions, and auxiliary proteins. In addition, we have preliminary data showing that delta receptor function is dependent on connections with trans-synaptic protein complexes, therefore we will study the structure and function of delta receptors in the context of synaptic binding partners. Together, these investigations will place our fundamental single-molecule investigations in a physiological context, and eventually allow us to understand functional differences between synaptic and non-synaptic glutamate receptors.