Crystal Structure of the Photosensory Module from a PAS-Less Cyanobacterial Phytochrome as Pr Shows a Mix of Dark-Adapted and Photoactivated Features

E Sethe Burgie; Alayna J Mickles; Fang Luo; Mitchell D Miller; Richard D Vierstra

doi:10.1016/j.jbc.2024.107369

Crystal Structure of the Photosensory Module from a PAS-Less Cyanobacterial Phytochrome as Pr Shows a Mix of Dark-Adapted and Photoactivated Features

J Biol Chem. 2024 May 13:107369. doi: 10.1016/j.jbc.2024.107369. Online ahead of print.

Authors

E Sethe Burgie¹, Alayna J Mickles¹, Fang Luo¹, Mitchell D Miller², Richard D Vierstra³

Affiliations

¹ Department of Biology, Washington University in St. Louis, St. Louis, Missouri 63130 USA.
² Department of Biosciences, Rice University, Houston, Texas 77005 USA.
³ Department of Biology, Washington University in St. Louis, St. Louis, Missouri 63130 USA. Electronic address: rdvierstra@wustl.edu.

PMID: 38750792
DOI: 10.1016/j.jbc.2024.107369

Abstract

Phytochromes (Phys) are a diverse collection of photoreceptors that regulate numerous physiological and developmental processes in microorganisms and plants through photointerconversion between red-light absorbing Pr and far-red light-absorbing Pfr states. Light is detected by an N-terminal photosensing module (PSM) sequentially comprised of Period/ARNT/Sim (PAS), cGMP-phosphodiesterase/adenylyl cyclase/FhlA (GAF), and Phy-specific (PHY) domains, with the bilin chromophore covalently-bound within the GAF domain. Phys sense light via the Pr/Pfr ratio measured by light-induced rotation of the bilin D-pyrrole ring that triggers conformational changes within the PSM, which for microbial Phys reaches into an output region. A key step is a β-stranded to α-helical reconfiguration of a hairpin loop extending from the PHY domain to contact the GAF domain. Besides canonical Phys, cyanobacteria express several variants, including a PAS-less subfamily that harbors just the GAF and PHY domains for light detection. Prior 2D-NMR studies of a model PAS-less Phy from Synechococcus_sp._JA-2-3B'a(2-13) (SyB-Cph1) proposed a unique photoconversion mechanism involving an A-pyrrole ring rotation, while magic-angle-spinning NMR probing the chromophore proposed the prototypic D-ring flip. To help solve this conundrum, we determined the crystallographic structure of the GAF-PHY region from SyB-Cph1 as Pr. Surprisingly, this structure differs from canonical Phys by having a Pr ZZZsyn,syn,anti bilin configuration but shifted to the activated position in the binding pocket with consequent folding of the hairpin loop to α-helical, an architecture common for Pfr. Collectively, the PSM of SyB-Cph1 as Pr displayed a mix of dark-adapted and photoactivated features whose co-planar A-C pyrrole rings supports a D-ring flip mechanism.

Keywords: PAS-less Phy; Synechococcus sp. JA-2-3B'a(2-13); X-ray crystallography; bilin chromophore; cyanobacteria; phytochrome; red light-absorbing Pr state.