Structural analysis of the regulatory GAF domains of cGMP phosphodiesterase elucidates the allosteric communication pathway

Authors

Richa Gupta, University of New Hampshire, Durham
Yong Liu, University of North Carolina at Chapel Hill
Huanchen Wang, Signal Transduction Laboratory, NIEHS/NIH
Christopher T. Nordyke, University of New Hampshire, Durham
Ryan Z. Puterbaugh, University of New Hampshire, Durham
Wenjin Cui, University of New Hampshire, Durham
Krisztina Varga, University of New Hampshire, Durham
Feixia Chu, University of New Hampshire, Durham
Hengming Ke, University of New Hampshire, Durham
Harish Vashisth, University of New Hampshire, Durham
Rick H. Cote, University of New Hampshire - Main CampusFollow

Abstract

Regulation of photoreceptor phosphodiesterase (PDE6) activity is responsible for the speed, sensitivity, and recovery of the photoresponse during visual signaling in vertebrate photoreceptor cells. It is hypothesized that the physiological differences in the light responsiveness of rods and cones may result in part from differences in the structure and regulation of the distinct isoforms of rod and cone PDE6. Although rod and cone PDE6 catalytic subunits share a similar domain organization consisting of tandem GAF domains (GAFa and GAFb) and a catalytic domain, cone PDE6 is a homodimer whereas the rod PDE6 catalytic dimer is composed of two homologous catalytic subunits. Here we provide the x-ray crystal structure of cone GAFab regulatory domain solved at 3.3 Å resolution in conjunction with chemical cross-linking and mass spectrometric analysis of conformational changes to GAFab induced upon binding of cGMP and the PDE6 inhibitory γ-subunit (Pγ). Ligand-induced changes in cross-linked residues implicate the α4-helix of GAFa (close to the cGMP binding site) and the β1/β2 loop of GAFb as key motifs that have been previously proposed to communicate with the catalytic domains of PDE6. Molecular dynamics (MD) simulations of cone GAFab revealed asymmetry in the two GAFab subunits forming the homodimer and allosteric perturbations on cGMP binding. Cross-linking of Pγ to GAFab in conjunction with solution NMR spectroscopy of isotopically labeled Pγ identified the central polycationic region of Pγ interacting with the GAFb domain. These results provide a mechanistic basis for developing allosteric activators of PDE6 with therapeutic implications for halting the progression of certain retinal degenerative diseases.

Department

Molecular, Cellular and Biomedical Sciences

Publication Date

11-18-2019

Digital Object Identifier (DOI)

https://dx.doi.org/10.34051/p/2019.5

Document Type

Article

Recommended Citation

Gupta, Richa; Liu, Yong; Wang, Huanchen; Nordyke, Christopher T.; Puterbaugh, Ryan Z.; Cui, Wenjin; Varga, Krisztina; Chu, Feixia; Ke, Hengming; Vashisth, Harish; and Cote, Rick H., "Structural analysis of the regulatory GAF domains of cGMP phosphodiesterase elucidates the allosteric communication pathway" (2019). Faculty Publications. 705.
https://scholars.unh.edu/faculty_pubs/705