Activation of the retinal cGMP-specific phosphodiesterase by the GDP-loaded alpha-subunit of transducin.
The interaction of the GDP-bound form of the alpha-subunit of transducin (T alpha GDP) with the cGMP-specific phosphodiesterase, the effector enzyme in the visual system, has been studied. T alpha GDP is demonstrated to be able to activate the phosphodiesterase: (a) the basal activity in suspensions of dark-adapted retinal rod outer segments, examined in the absence of GTP, was found to be inhibited by binding of transducin to activated rhodopsin (Rh*) and by the complex of the beta- and gamma-subunits of transducin (T beta gamma); (b) purified T alpha GDP is able to activate phosphodiesterase in the presence of membranes; (c) no activation is obtained either with holotransducin (T alpha GDP T beta gamma) or with T alpha GDP in the presence of excess T beta gamma to prevent dissociation of TGDP. The maximal level of phosphodiesterase activation reached with T alpha GDP (about 1500 mol cGMP/mol phosphodiesterase-1.s-1) is similar to that obtained through the 'classical' activation by T alpha GTP whereas the apparent affinity of T alpha GDP for phosphodiesterase (Kd about 50 microM) is much lower than that of T alpha GTP. Our data suggest that GTP hydrolysis itself does not inactivate T alpha. The role of T beta gamma to sequester T alpha is therefore of critical importance for phosphodiesterase inactivation. Our results support observations on the regulation of adenylyl cyclase by G-proteins, which suggested the ability of the free alpha-subunits loaded with GDP to activate their effectors.
Kutuzov
M
M
Département de Biologie Moléculaire et Structurale, Centre d'Etudes Nucléaires, Grenoble, France.
Pfister
C
C
D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000378:metabolism*; D000818:Animals; D002417:Cattle; D003624:Darkness; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000378:metabolism; D006153:Guanosine Diphosphate / Q000378:metabolism; D066298:In Vitro Techniques; D007425:Intracellular Membranes / Q000201:enzymology; D012374:Rod Cell Outer Segment / Q000201:enzymology*; D015293:Transducin / Q000378:metabolism*
D006153:Guanosine Diphosphate; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D019204:GTP-Binding Proteins; D015293:Transducin