title: Protonation-Dependent Structural Heterogeneity in the Chromophore Binding Site of Cyanobacterial Phytochrome Cph1.
abstract: Phytochromes are biological red/far-red light sensors found in many organisms. Photoisomerization of the linear methine-bridged tetrapyrrole triggers transient proton translocation events in the chromophore binding pocket (CBP) leading to major conformational changes of the protein matrix that are in turn associated with signaling. By combining pH-dependent resonance Raman and UV-visible absorption spectroscopy, we analyzed protonation-dependent equilibria in the CBP of Cph1 involving the proposed Pr-I and Pr-II substates that prevail below and above pH 7.5, respectively. The protonation pattern and vibrational properties of these states were further characterized by means of hybrid quantum mechanics/molecular mechanics calculations. From this combined experimental-theoretical study, we were able to identify His260 as the key residue controlling pH-dependent equilibria. This residue is not only responsible for the conformational heterogeneity of CBP in the Pr state of prokaryotic phytochromes, discussed extensively in the past, but it constitutes the sink and source of protons in the proton release/uptake mechanism involving the tetrapyrrole chromophore which finally leads to the formation of the Pfr state. Thus, this work provides valuable information that may guide further experiments toward the understanding of the specific role of protons in controlling structure and function of phytochromes in general.
authors.affiliation: Institut für Chemie, Technische Universität Berlin , Sekr. PC 14, Straße des 17. Juni 135, D-10623 Berlin, Germany.
authors.forename: Francisco
authors.initials: F
authors.lastname: Velazquez Escobar
authors.affiliation: Plant Physiology, Justus-Liebig University Gießen , Senckenbergstrasse 3, D-35390 Giessen, Germany.
authors.forename: Christina
authors.initials: C
authors.lastname: Lang
authors.affiliation: Plant Physiology, Justus-Liebig University Gießen , Senckenbergstrasse 3, D-35390 Giessen, Germany.
authors.forename: Aref
authors.initials: A
authors.lastname: Takiden
authors.affiliation: Institut für Experimentalphysik, Freie Universität Berlin , Arnimallee 14, 14195 Berlin, Germany.
authors.forename: Constantin
authors.initials: C
authors.lastname: Schneider
authors.affiliation: Institut für Experimentalphysik, Freie Universität Berlin , Arnimallee 14, 14195 Berlin, Germany.
authors.forename: Jens
authors.initials: J
authors.lastname: Balke
authors.affiliation: Plant Physiology, Justus-Liebig University Gießen , Senckenbergstrasse 3, D-35390 Giessen, Germany.
authors.forename: Jon
authors.initials: J
authors.lastname: Hughes
authors.affiliation: Institut für Experimentalphysik, Freie Universität Berlin , Arnimallee 14, 14195 Berlin, Germany.
authors.forename: Ulrike
authors.initials: U
authors.lastname: Alexiev
authors.affiliation: Institut für Chemie, Technische Universität Berlin , Sekr. PC 14, Straße des 17. Juni 135, D-10623 Berlin, Germany.
authors.forename: Peter
authors.identifier: 0000-0003-1030-5900
authors.initials: P
authors.lastname: Hildebrandt
authors.affiliation: Institut für Chemie, Technische Universität Berlin , Sekr. PC 14, Straße des 17. Juni 135, D-10623 Berlin, Germany.
authors.forename: Maria Andrea
authors.identifier: 0000-0002-7497-5631
authors.initials: MA
authors.lastname: Mroginski
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry*; D001665:Binding Sites; D000458:Cyanobacteria / Q000737:chemistry*; D020283:Photoreceptors, Microbial; D010834:Phytochrome / Q000737:chemistry*; D011487:Protein Conformation; D011494:Protein Kinases / Q000737:chemistry*; D011522:Protons; D011789:Quantum Theory
chemical_list: D001426:Bacterial Proteins; D020283:Photoreceptors, Microbial; D011522:Protons; D010834:Phytochrome; D011494:Protein Kinases; C413623:Cph1 phytochrome protein, bacteria