Solid‐State NMR Spectroscopy to Probe Photoactivation in Canonical Phytochromes

Abstract

The photoreceptor phytochrome switches photochromically between two thermally stable states called Pr and Pfr. Here, we summarize recent solid-state magic-angle spinning (MAS) NMR work on this conversion process and interpret the functional mechanism in terms of a nano-machine. The process is initiated by a double-bond photoisomerization of the open-chain tetrapyrrole chromophore at the methine bridge connecting pyrrole rings C and D. The Pr-state chromophore and its surrounding pocket in canonical cyanobacterial and plant phytochromes has significantly less order, tends to form isoforms and is soft. Conversely, Pfr shows significantly harder chromophore-protein interactions, a well-defined protonic and charge distribution with a clear classical counterion for the positively charged tetrapyrrole system. The soft-tohard/disorder-to-order transition involves the chromophore and its protein surroundings within a sphere of at least 5.5 A. The relevance of this collective event for signaling is discussed. Measurement of the intermediates during the Pfr ? Pr back-reaction provides insight into the well-adjusted mechanics of a two-step transformation. As both Pr ? Pfr and Pfr ? Pr reaction pathways are different in ground and excited states, a photochemically controlled hyper-landscape is proposed allowing for ratchet-type reaction dynamics regulating signaling activity.

New Constitutively Active Phytochromes Exhibit Light-Independent Signaling Activity

“…They are biosynthesized as the Pr form in the dark, which can be phototransformed into the biologically active Pfr form upon exposure to R light. The absorption of R light by the Pr form triggers a Z-to-E isomerization of the chromophore and leads to reversible conformational changes throughout the protein moiety, resulting in the Pfr form (Vierstra and Zhang, 2011;Song et al, 2013;. Conversely, the Pfr form can be converted to the Pr form by the absorption of FR light.…”

Structure of the Biliverdin Cofactor in the Pfr State of Bathy and Prototypical Phytochromes

“…In this context it is interesting to refer to recent NMR studies of the prototypical cyanobacterial phytochrome Cph1 (65). These authors have shown a less ordered chromophore pocket in the Pr state, whereas in the Pfr state the chromophore gives rise to well defined resonances with no indication for heterogeneities.…”

The D-ring, Not the A-ring, Rotates in Synechococcus OS-B′ Phytochrome

“…In general, red light absorption by the Pr 2 dark state leads to formation of the Pfr state that is photoconverted by far-red light back to Pr, physiological signaling being switched by state transition. The structural characteristics of Pr and Pfr, the photoconversion mechanism, and the physiological signals generated are all subjects of intense research (1)(2)(3).…”