Quaternary organization of a phytochrome dimer as revealed by cryoelectron microscopy

Abstract

Phytochromes are a collection of dimeric photoreceptors that direct a diverse array of responses in plants and microorganisms through photoconversion between a red light-absorbing ground state Pr, and a far-red light-absorbing photoactivated state Pfr. Photoconversion from Pr to Pfr is initiated by a light-driven rotation within the covalently attached bilin, which then triggers a series of protein conformational changes in the binding pocket. These movements ultimately affect an appended output module, which often has reversible protein kinase activity. Propagation of the light signal from the bilin to the output module likely depends on the dimerization interface but its architecture and response to phototransformation remain unclear. Here, we used single particle cryoelectron microscopy to determine the quaternary arrangement of the phytochrome dimer as Pr, using the bacteriophytochrome (BphP) from Deinococcus radiodurans . Contrary to the long-standing view that the two monomers are held together solely via their C-terminal region, we provide unambiguous evidence that the N-terminal bilin-binding region of BphP also provides a dimerization interface with the C-terminal kinase domain appearing as a more flexible appendage. The BphP monomers dimerize in parallel with the polypeptides intimately twisting around each other in a right-handed fashion. Based on this electron microscopic picture, we propose that the light-driven conformational changes transmitted from the chromophore to the output module along the spine of this extensive dimer interface is the central feature underpinning phytochrome signaling.

Structure of the Cyanobacterial Phytochrome 2 Photosensor Implies a Tryptophan Switch for Phytochrome Signaling

“…Its staggered and antiparallel orientation differs from other known phytochrome structures such as PaBPhP, in which a parallel association is observed over the whole length of the GAF-PHY linker helices. The latter mode of association is thought to be crucial for signal transduction to histidine kinase domains (9,36) and illustrated by the cryoEM structure of fulllength bacteriophytochrome DrBphP, where the photosensory module provides the largest dimerization interface. Like in parallel dimers of the photosensory phytochrome modules, the antiparallel arrangement of SynCph2 (1-2) is mainly stabilized by association of its linker ␣ 5 -helices between GAF1 and GAF2 (Figs.…”

Crystal structure of the photosensing module from a red/far-red light-absorbing plant phytochrome

“…SEC supported an elongated shape for the crystallographic subunits by estimating a Stokes radius ∼1.3 times that predicted. Analogous to several BphPs (12,13,17), the dimerization interface involved the GAF domain α1/α2/α6-helical bundle ( Fig. 1A and Fig.…”

Phytochromes: An Atomic Perspective on Photoactivation and Signaling

“…Included are x-ray crystallographic and/or solution 2D-NMR structures of the GAF domain from PAS-less Phys Anders et al, 2013), a Phy that photoconverts from Pr to the Pnr state (Yang et al, 2007), and CBCRs (Burgie et al, 2013;Narikawa et al, 2013;Cornilescu et al, 2014), crystal structures of the entire PSM region (PAS-GAF-PHY) from canonical Phys (Essen et al, 2008;Burgie et al, 2014b;Takala et al, 2014) and bathyPhys (Yang et al, 2008;Bellini and Papiz, 2012), and even images of entire Phy dimers by small-angle x-ray scattering (Evans et al, 2006) and single-particle electron microscopy (SPEM) (Li et al, 2010). Particularly informative have been paired structures of the dark-adapted and photoactivated states (Ulijasz et al, 2010;Cornilescu et al, 2014;Takala et al, 2014), comparisons of canonical and bathy-Phys (Yang et al, 2009), and solid-state NMR analyses of the bilin and temperature-scanning cryocrystallography following sample irradiation (Song et al, 2011;Yang et al, 2011) that have collectively illuminated the structural changes associated with photointerconversion.…”