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Molecular architecture of photoreceptor phosphodiesterase (PDE6) with activated G protein elucidates the mechanism of visual excitation

ABSTRACT: Photoreceptor phosphodiesterase 6 (PDE6) is the central effector of the visual excitation pathway in both rod and cone photoreceptors, and PDE6 mutations that alter PDE6 structure or regulation can result in several human retinal diseases. The rod PDE6 holoenzyme consists of two catalytic subunits (Pαβ) whose activity is suppressed in the dark by binding of two inhibitory γ-subunits (Pγ). Upon photoactivation of rhodopsin, the heterotrimeric G protein (transducin)is activated, resulting in binding of the activated transducin α-subunit (Gtα) toPDE6, displacement of Pγ from the PDE6 active site, and enzyme activation. Although the biochemistry of this pathway is understood, a lack of detailed structural information about the PDE6 activation mechanism hampers efforts to develop therapeutic interventions for managing PDE6-associated retinal disease. To address this gap, here we used a cross-linking MS-based approach to create a model of the entire interaction surface of Pγ with the regulatory and catalytic domains of Pαβin its nonactivated state. Following reconstitution of PDE6 and activated Gtαwith liposomes and identification of cross-links between Gtα and PDE6 subunits, we determined that the PDE6-Gtα protein complex consists of two Gtα binding sites per holoenzyme. Each Gtα interacts with the catalytic domains of both catalytic subunits and induces major changes in the interaction sites of interaction of the Pγ subunit with the catalytic subunits. These results provide the first structural model for the activated state of the transducin-PDE6-Tα*complex during visual excitation, thereby enhancing our understanding of the molecular etiology of inherited retinal diseases.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Bos Taurus (bovine)

TISSUE(S): Retinal Rod

SUBMITTER: Rick Cote

LAB HEAD: Rick H. Cote

PROVIDER: PXD015989 | Pride | 2019-11-11

REPOSITORIES: Pride

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The molecular architecture of photoreceptor phosphodiesterase 6 (PDE6) with activated G protein elucidates the mechanism of visual excitation.

Irwin Michael J MJ Gupta Richa R Gao Xiong-Zhuo XZ Cahill Karyn B KB Chu Feixia F Cote Rick H RH

The Journal of biological chemistry 20191105 51

Photoreceptor phosphodiesterase 6 (PDE6) is the central effector of the visual excitation pathway in both rod and cone photoreceptors, and <i>PDE6</i> mutations that alter PDE6 structure or regulation can result in several human retinal diseases. The rod PDE6 holoenzyme consists of two catalytic subunits (Pαβ) whose activity is suppressed in the dark by binding of two inhibitory γ-subunits (Pγ). Upon photoactivation of rhodopsin, the heterotrimeric G protein (transducin) is activated, resulting ...[more]

PMID: 31690623

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