The goal of the proposed research is to elucidate the molecular mechanism of visual excitation in vertebrate retinal rods. Phototransduction is now known to involve a light-activated enzyme cascade that leads to a rapid reduction of cytosolic cycli GMP and the closure of Na+ channels at the plasma membranes. Central to this process is the activation of three proteins: transducin, cyclic GMP phosphodiesterase, and a postulated cyclic GMP-sensitive channel. The goal of this study is to gain and understanding of the molecular basis of photoexcitation through detailed strucutral and functional analyses of these retinal proteins. We propose to carry out the following studies: (1) Specific monoclonal antibody probes directed against transducin and phosphodiesterase will be used to investigate the structure-function relationships of these proteins, to localize their distributions in photoreceptors, and to study the cause of retinal degenerations. (2) Site-directed chemical probes and anti-synthetic peptdie antibodies will be developed to identify and study the active sites of transducin, and to characterize other cellular GPT-binding homologs. (3) The interaction of the lambda inhibitor and cyclic GMP with phosphodiesterase will be invesigated. The primary structure of the phosphodiesterase subunits will be elucidated by recombinant DNA techniques. (4) Finally, an attempt will be made to isolate and identify the cyclic GMP-sensitive channel. By advancing our knowledge in these four specific areas, our research is expected to contribute to the overall goal of achieving a better understanding of how the interactions of these proteins give rise to activation and termination of the photoresponse in visual cells.
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