title: Conformational heterogeneity of the Pfr chromophore in plant and cyanobacterial phytochromes.
abstract: Phytochromes are biological photoreceptors that can be reversibly photoconverted between a dark and photoactivated state. The underlying reaction sequences are initiated by the photoisomerization of the tetrapyrrole cofactor, which in plant and cyanobacterial phytochromes are a phytochromobilin (PΦB) and a phycocyanobilin (PCB), respectively. The transition between the two states represents an on/off-switch of the output module activating or deactivating downstream physiological processes. In addition, the photoactivated state, i.e., Pfr in canonical phytochromes, can be thermally reverted to the dark state (Pr). The present study aimed to improve our understanding of the specific reactivity of various PΦB- and PCB-binding phytochromes in the Pfr state by analysing the cofactor structure by vibrational spectroscopic techniques. Resonance Raman (RR) spectroscopy revealed two Pfr conformers (Pfr-I and Pfr-II) forming a temperature-dependent conformational equilibrium. The two sub-states-found in all phytochromes studied, albeit with different relative contributions-differ in structural details of the C-D and A-B methine bridges. In the Pfr-I sub-state the torsion between the rings C and D is larger by ca. 10° compared to Pfr-II. This structural difference is presumably related to different hydrogen bonding interactions of ring D as revealed by time-resolved IR spectroscopic studies of the cyanobacterial phytochrome Cph1. The transitions between the two sub-states are evidently too fast (i.e., nanosecond time scale) to be resolved by NMR spectroscopy which could not detect a structural heterogeneity of the chromophore in Pfr. The implications of the present findings for the dark reversion of the Pfr state are discussed.
authors.affiliation: Institut für Chemie, Technische Universität Berlin Berlin, Germany.
authors.forename: Francisco
authors.initials: F
authors.lastname: Velazquez Escobar
authors.affiliation: Institut für Chemie, Technische Universität Berlin Berlin, Germany.
authors.forename: David
authors.initials: D
authors.lastname: von Stetten
authors.affiliation: Institut für Chemie, Technische Universität Berlin Berlin, Germany.
authors.forename: Mina
authors.initials: M
authors.lastname: Günther-Lütkens
authors.affiliation: Institut für Chemie, Technische Universität Berlin Berlin, Germany.
authors.forename: Anke
authors.initials: A
authors.lastname: Keidel
authors.affiliation: Institut für Chemie, Technische Universität Berlin Berlin, Germany.
authors.forename: Norbert
authors.initials: N
authors.lastname: Michael
authors.affiliation: Botanisches Institut, Karlsruher Institut für Technologie Karlsruhe, Germany.
authors.forename: Tilman
authors.initials: T
authors.lastname: Lamparter
authors.affiliation: Fachbereich Chemie, Philipps-Universität Marburg Marburg, Germany.
authors.forename: Lars-Oliver
authors.initials: LO
authors.lastname: Essen
authors.affiliation: Institut für Pflanzenphysiologie, Justus Liebig University Gießen, Germany.
authors.forename: Jon
authors.initials: J
authors.lastname: Hughes
authors.affiliation: Max-Planck-Institut für Chemische Energiekonversion Mülheim, Germany.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Gärtner
authors.affiliation: Institut für Experimentalphysik, Freie Universität Berlin Berlin, Germany.
authors.forename: Yang
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Institut für Experimentalphysik, Freie Universität Berlin Berlin, Germany.
authors.forename: Karsten
authors.initials: K
authors.lastname: Heyne
authors.affiliation: Institut für Chemie, Technische Universität Berlin Berlin, Germany.
authors.forename: Maria A
authors.initials: MA
authors.lastname: Mroginski
authors.affiliation: Institut für Chemie, Technische Universität Berlin Berlin, Germany.
authors.forename: Peter
authors.initials: P
authors.lastname: Hildebrandt
keywords: hydrogen bonding; isomerization; phytochrome; quantum chemical calculations; resonance Raman spectroscopy; structural heterogeneity; tetrapyrrole; time-resolved IR spectroscopy