Possible Stimulation of Retinal Rod Recovery to Dark State by cGMP Release from a cGMP Phosphodiesterase Noncatalytic Site

• Published in: The Journal of biological chemistry Vol. 271; no. 51; pp. 32495 - 32498
• Main Authors: Yamazaki, Akio, Bondarenko, Vladimir A., Dua, Salil, Yamazaki, Matsuyo, Usukura, Jiro, Hayashi, Fumio
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 20.12.1996; American Society for Biochemistry and Molecular Biology
• Subjects: 3',5'-Cyclic-GMP Phosphodiesterases - metabolism; Animals; Binding Sites; Cell Membrane - metabolism; Cyclic GMP - metabolism; Darkness; Mutagenesis, Site-Directed; Photoreceptor Cells - metabolism; Rana catesbeiana; Structure-Activity Relationship; Vision, Ocular
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.271.51.32495

Cyclic GMP phosphodiesterase, a key enzyme for phototransduction, contains α, β (Pαβ), and two γ (Pγ) subunits. In addition to catalytic sites, Pαβ has two classes of noncatalytic cGMP binding sites with different affinities (Kd values <100 nM and >1 μM). Pγ regulates Pαβ as an inhibitor of cGMP hydrolysis and as a stimulator of cGMP binding to the high affinity noncatalytic sites. Pγ release from Pαβ by the GTP-bound α subunit of transducin (GTP·Tα) interrupts these two functions. Here we describe a novel regulation of the Pγ release by [cGMP] and its physiological implication. We isolated Pγ mutants that exhibit abnormally one of these two functions, indicating the distinct domains in Pγ are involved to express these functions. When [cGMP] was high (~5 μM), Pγ responsible for the inhibition of cGMP hydrolysis was preferentially released, and cGMP hydrolysis activity of Pαβ was increased about 10 times. When [cGMP] was low (less than ~0.5 μM), Pγ responsible for the stimulation of cGMP binding to the high affinity sites was released. The Pγ release resulted in the decrease of relative affinity of cGMP for the high affinity sites to at least (null)/1;10, followed by the rapid release of cGMP from one of the high affinity sites (apparent t1/2 = 3.8 s). cGMP (~5 μM) inhibited the extraction of Pαβ from rod membranes by a Mg2+-free hypotonic buffer. The inhibition of Pαβ extraction was not affected by Pγ, suggesting that Pαβ detects on the order of micromolar [cGMP] using low affinity noncatalytic sites on Pαβ. Because [cGMP] is ~5 μM in darkness and lowered by photoexcitation and phosphodiesterase concentration is ~30 μM in rod photoreceptors, it is possible that cGMP phosphodiesterase functions to increase cytoplasamic [cGMP] after [cGMP] is reduced to the illuminated level.