Functional mapping of interacting regions of the photoreceptor phosphodiesterase (PDE6) γ-subunit with PDE6 catalytic dimer, transducin, and regulator of G-protein signaling9-1 (RGS9-1).
The cGMP phosphodiesterase (PDE6) involved in visual transduction in photoreceptor cells contains two inhibitory γ-subunits (Pγ) which bind to the catalytic core (Pαβ) to inhibit catalysis and stimulate cGMP binding to the GAF domains of Pαβ. During visual excitation, interaction of activated transducin with Pγ relieves inhibition. Pγ also participates in a complex with RGS9-1 and other proteins to accelerate the GTPase activity of activated transducin. We studied the structural determinants for these important functions of Pγ. First, we identified two important sites in the middle region of Pγ (amino acids 27-38 and 52-54) that significantly stabilize the overall binding affinity of Pγ with Pαβ. The ability of Pγ to stimulate noncatalytic cGMP binding to the GAF domains of PDE6 has been localized to amino acids 27-30 of Pγ. Transducin activation of PDE6 catalysis critically depends on the presence of Ile54 in the glycine-rich region of Pγ in order to relieve inhibition of catalysis. The central glycine-rich region of Pγ is also required for transducin to increase cGMP exchange at the GAF domains. Finally, Thr-65 and/or Val-66 of Pγ are critical residues for Pγ to stimulate GTPase activity of transducin in a complex with RGS9-1. We propose that the glycine-rich region of Pγ is a primary docking site for PDE6-interacting proteins involved in the activation/inactivation pathways of visual transduction. This functional mapping of Pγ with its binding partners demonstrates the remarkable versatility of this multifunctional protein and its central role in regulating the activation and lifetime of visual transduction.
Department of Molecular, Cellular, and Biomedical Sciences, University of New Hampshire, Durham, New Hampshire 03824, USA.
Xiu-Jun
XJ
Zhang
Xiong-Zhuo
XZ
Gao
Wei
W
Yao
Rick H
RH
Cote
D000818:Animals; D001665:Binding Sites; D002417:Cattle; D006152:Cyclic GMP / Q000737:chemistry; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003553:Cystine / Q000737:chemistry; D005136:Eye Proteins / Q000737:chemistry / Q000378:metabolism; D007532:Isoleucine / Q000737:chemistry; D010446:Peptide Fragments / Q000737:chemistry; D010449:Peptide Mapping; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D021122:Protein Subunits / Q000737:chemistry; D020710:RGS Proteins / Q000737:chemistry* / Q000378:metabolism; D055214:Retinal Photoreceptor Cell Outer Segment / Q000201:enzymology / Q000378:metabolism; D015293:Transducin / Q000737:chemistry* / Q000378:metabolism
D005136:Eye Proteins; D010446:Peptide Fragments; D021122:Protein Subunits; D020710:RGS Proteins; D007532:Isoleucine; D003553:Cystine; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6; D015293:Transducin; D006152:Cyclic GMP