Tertiary Structures of Potassium Channel Domains

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DESCRIPTION (provided by applicant): This is a competitive continuation proposal to support three new aims of the project entitled "Tertiary structures of potassium channel domains." This renewal proposal expands the scope and direction to a newer level by focusing on molecular working of 6 transmembrane (TM)-helix voltage-gated K channels. In the first aim, we propose to determine a bacterial 6 TM K channel, KvPae in complex with an FLAG-directed Fab complex. This work will address the structural basis of voltage-mediated channel activation and its mechanism, and conformational flexibility endowed in the channel. In the second aim, we propose to complete the structure of the entire N-terminal domain of eukaryotic Kv channel, aKv1.1, and analyze the conformational change by NMR spectroscopy combined with functional characterization. This work will address the structural basis of the mechanism of channel inactivation by its own inactivation subdomain by protein-protein interaction. In the third aim, we propose to study structural changes occurring in the complex of calcium-binding cytoplasmic KChIP protein in complex with the "entire" N-terminal domain of eukaryotic Kv channel, Kv4.2. Calcium-induced conformational change in KChIP/channel complex will be probed by a combination of x-ray crystallography and NMR spectroscopy methods. This study will expand our molecular understanding of diverse regulatory mechanisms of eukaryotic potassium channels. We will learn a great deal about structural mechanisms underlying voltage-activation (Aim 1), inactivation mediated by its subdomain (Aim 2), and calcium-mediated regulation (Aim 3).