Activation mechanism of rod outer segment cyclic GMP phosphodiesterase. Release of inhibitor by the GTP/GTP-binding protein.
The physiological regulation of light-activated cyclic GMP phosphodiesterase (EC 3.1.4.17) in rod outer segments has been shown to depend upon a heat-stable inhibitor and upon the reversal of its effect by a specific GTP/GTP-binding protein complex (Hurley, J. B. (1980) Biochem. Biophys. Res. Commun. 92, 505-510; Yamazaki, A., Bartucca, F., Ting, A., and Bitensky, M. W. (1982) Proc. Natl. Acad. Sci. U. S. A. 79, 3702-3706). Washing of illuminated disc membranes with an isotonic buffer released 86% of the peripheral proteins without any release of inhibitor. Subsequent washing with the same isotonic buffer containing GTP released 80% of the inhibitor. When inhibitor was eluted with guanosine-5'-(beta, gamma-imino)triphosphate, it had an apparent molecular weight of 60,000 on Sephadex G-100. The release of inhibitor by guanosine-5'-(beta, gamma-imino)triphosphate was also demonstrated with sucrose density gradient centrifugation. Inhibitor release from the disc membrane by GTP or its analogue was accompanied by the release of the GTP-binding protein and an increased phosphodiesterase activity in the membrane. However, following GTP hydrolysis, both inhibitor and GTP-binding protein returned to the membrane and phosphodiesterase activity in the membrane decreased proportionally. In contrast, incubation of disc membranes with guanosine-5'-(beta, gamma-imino)-triphosphate produced an increase of inhibitor activity in the supernatant and an increase of phosphodiesterase activity in the pellet which remained constant after the initial increase. These data clearly show that the activation of phosphodiesterase by the GTP/GTP-binding protein complex resulted from the release of inhibitor. Hydrolysis of GTP resulted in the reassociation of inhibitor with and concomitant inhibition of disc membrane phosphodiesterase.
D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000378:metabolism*; D000818:Animals; D019204:GTP-Binding Proteins; D006160:Guanosine Triphosphate / Q000378:metabolism*; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D007700:Kinetics; D008027:Light; D008970:Molecular Weight; D010786:Photoreceptor Cells / Q000201:enzymology*; D011892:Rana catesbeiana; D011956:Receptors, Cell Surface / Q000302:isolation & purification / Q000378:metabolism*; D012374:Rod Cell Outer Segment / Q000201:enzymology*
D011956:Receptors, Cell Surface; D006165:Guanylyl Imidodiphosphate; D006160:Guanosine Triphosphate; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D019204:GTP-Binding Proteins