Single and multi-channel conductance measurements, high resolution structural and dynamic characterization by solid-state NMR, and high level computational efforts will lead to structure-function and dynamic-function correlations. These will greatly facilitate our understanding of ion channel conductance, gating and blockage mechanisms, as well as numerous functional details. Previous support through this grant has led to novel correlations for defining functional mechanisms, ion specificity and efficiency in gramicidin A, as well as unique models for opening and closing this channel. Initial mechanistic insights have also been obtained for the M2 H+ channel from influenza A virus. Electrophysiological and NMR studies of the full length M2 protein have illustrated how sensitive membrane proteins are to their bilayer environment and how these structures can have dynamic processes rarely, if ever, observed in water soluble proteins. ? ? Here, we propose to characterize structure-dynamic-function correlations in the M2 H+ channel leading to mechanistic understandings. This protein is a proven drug target and it is the first H+ channel to be cloned and expressed. Several key features of this structure include the single transmembrane helix that can be kinked in the presence of a channel blocker, a histidine tetrad that appears to be responsible for H+ selectivity and acid gating, and a tryptophan tetrad that interacts with the histidines, at least in the closed state. High resolution backbone structure and dynamics, as a function of pH, will be obtained for this protein, as well as the pKas of the histidines under various conditions. These efforts will be guided by both modeling and functional characterizations of the protein. ? ? This work has broad implications for understanding how, in general, membrane proteins function and how ion specificity, conductance efficiency, gating and blockage can be achieved and characterized all in a lamellar phase lipid environment. ? ?
Showing the most recent 10 out of 99 publications
