Structure-function studies on Ion channels

Project Summary: Our research program investigates the functional mechanisms that operate in voltage gated K+ (Kv) channels. Kv channels are essential for the generation and conduction of electrical signals by neurons, muscles and endocrine cells. Underlying the physiological roles of Kv channels is their ability to regulate the flux of K+ across cellular membranes. The flux of K+ through a Kv channel is controlled by the processes of voltage activation, which turns on the flux, and inactivation, which turns off the flux of K+ through the channel. There have been extensive investigations on Kv channels but major questions remain on the activation and inactivation mechanisms. In our research program, we use a multidisciplinary approach that combines structural and functional studies with protein modifications using unnatural amino acid (UAA) mutagenesis. UAA mutagenesis is a very powerful method for protein modification compared to traditional mutagenesis because it allows a large variety of side chain modifications and permits the modification of the protein backbone. In the grant period, we will investigate C-type inactivation in the Shaker, Kv1.2 and the cardiac Kv channel HERG. C-type inactivation refers to a conformational change at the selectivity filter, the ion binding region of the channel, that turns off the flux of ions through the channel. We will also investigate the role of the protein main chain interactions in voltage gating. Our studies on voltage gating will be carried out in the hyperpolarization activated and cyclic nucleotide gated ion (HCN) channel. The research proposed is significant because it will provide key mechanistic insights into the working of Kv channels. The research will also provide strategies for investigating the role of the protein backbone and ion binding sites, strategies that will be generally applicable to other ion channels.