Light-induced activation of a cGMP phosphodiesterase (PDE) in rod outer segments of photoreceptors has been suggested to be a key step in the phototransduction process. This activation is mediated by a GTP-binding protein complex (G-protein). Although both the G-protein and the PDE have been purified from bovine and frog retina, to date, nothing is known of whether actual structural changes occur in these proteins during reginal degeneration. We will determine both quantitative and qualitative biochemical changes which occur in the ROS PDE from the rd mouse. The photoreceptors from the affected rd mice are deficient in a light-activated PDE activity resulting in high cGMP levels and retinal degeneration. We have found that the PDE protein is present in 10 day-old affected rd mice. However, it is not functional. Using recently developed techniques which are sensitive to less than ng levels we will 1) identify and quantitate normal and rd mouse PDE using western blotting and radioimmunoassays, 2) purify the proteins by immunoaffinity procedures, 3) characterize the biochemical changes which occur in these proteins during retinal degeneration using 2-dimensional peptide mapping, photoaffinity labelling, and antibody structural probing, and 4) elucidate the functional defects in the rd PDE using reconstitution systems. These data will provide basic knowledge of the structural and quantitative changes which occur in ROS disc membrane PDE during retinal degeneration in the rd mouse. Ultimately these data may aid in the development of drugs to control or prevent the degeneration of photoreceptor cells.
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