Voltage-sensitive ion channels form the molecular basis for electrical excitability of skeletal muscle, cardiac muscle, and the brain. In the heart, these channels play a major role in determining cardiac cell automaticity, excitability and conductivity. This proposal is aimed at elucidating the molecular basis of the structural and functional diversity of voltage-activated K+ channels, the structural basis of physiological properties, such as voltage sensitivity and ion selectivity and the regulation of a gene that encodes the delayed rectifier K+ channel. Site- directed mutagenesis will be used to identify and modify amino acid residues in the domain that senses and responds to changes in the membrane electrical field. Using cDNA cloning, the structure of different K+ channels will be compared and contrasted. The effects of structural modifications on channel activity and the activity of the newly cloned K+ channels will be assessed by voltage clamp and patch clamp recording from channels expressed in Xenopus oocytes or in stable transfection of muscle and nonmuscle cell lines. Furthermore, using the cDNA as a probe, we propose to characterize the gene for the rat skeletal muscle and cardiac delayed rectifier K+ channel. Site-directed antibodies will be used for the study of the assembly, membrane topology, and functional organization of the delayed rectifier K+ channel. Ion channels determine the susceptibility of the heart to arrhythmias. The studies proposed here are designed to achieve a new level of understanding, at the molecular level of structure-function relationship of K+ channels. The knowledge gained from these studies may lead to improved approaches to therapy of cardiac arrhythmias.
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