Molecular Architecture of Photoreceptor Phosphodiesterase Elucidated by Chemical Cross-Linking and Integrative Modeling

Photoreceptor phosphodiesterase (PDE6) is the central effector enzyme in visual excitation pathway in rod and cone photoreceptors. Its tight regulation is essential for the speed, sensitivity, recovery and adaptation of visual detection. Although major steps in the PDE6 activation/deactivation pathway have been identified, mechanistic understanding of PDE6 regulation is limited by the lack of knowledge about the molecular organization of the PDE6 holoenzyme (alpha beta gamma gamma). Here, we characterize the PDE6 holoenzyme by integrative structural determination of the PDE6 catalytic dimer (alpha beta), based primarily on chemical cross-linking and mass spectrometric analysis. Our models built from high-density cross-linking data elucidate a parallel organization of the two catalytic subunits, with juxtaposed alpha-helical segments within the tandem regulatory GAF domains to provide multiple sites for dimerization. The two catalytic domains exist in an open configuration when compared to the structure of PDE2 in the apostate. Detailed structural elements for differential binding of the gamma-subunit to the GAFa domains of the alpha- and beta-subunits are revealed, providing insight into the regulation of the PDE6 activation/deactivation cycle. (C) 2014 Elsevier Ltd. All rights reserved.