Institut für Chemie, Technische Universität Berlin, Sekr. PC 14, Strasse des 17. Juni 135, D-10623 Berlin, Germany.
Institut für Chemie, Technische Universität Berlin, Sekr. PC 14, Strasse des 17. Juni 135, D-10623 Berlin, Germany; Structural Biology Group, European Synchrotron Radiation Facility, 38043 Grenoble, France.
J Biol Chem. 2013 Jun 7;288(23):16800-16814. doi: 10.1074/jbc.M113.457531. Epub 2013 Apr 19.
Phytochromes act as photoswitches between the red- and far-red absorbing parent states of phytochromes (Pr and Pfr). Plant phytochromes display an additional thermal conversion route from the physiologically active Pfr to Pr. The same reaction pattern is found in prototypical biliverdin-binding bacteriophytochromes in contrast to the reverse thermal transformation in bathy bacteriophytochromes. However, the molecular origin of the different thermal stabilities of the Pfr states in prototypical and bathy bacteriophytochromes is not known. We analyzed the structures of the chromophore binding pockets in the Pfr states of various bathy and prototypical biliverdin-binding phytochromes using a combined spectroscopic-theoretical approach. For the Pfr state of the bathy phytochrome from Pseudomonas aeruginosa, the very good agreement between calculated and experimental Raman spectra of the biliverdin cofactor is in line with important conclusions of previous crystallographic analyses, particularly the ZZEssa configuration of the chromophore and its mode of covalent attachment to the protein. The highly homogeneous chromophore conformation seems to be a unique property of the Pfr states of bathy phytochromes. This is in sharp contrast to the Pfr states of prototypical phytochromes that display conformational equilibria between two sub-states exhibiting small structural differences at the terminal methine bridges A-B and C-D. These differences may mainly root in the interactions of the cofactor with the highly conserved Asp-194 that occur via its carboxylate function in bathy phytochromes. The weaker interactions via the carbonyl function in prototypical phytochromes may lead to a higher structural flexibility of the chromophore pocket opening a reaction channel for the thermal (ZZE → ZZZ) Pfr to Pr back-conversion.
光敏色素作为红-远红吸收母体状态的光开关（Pr 和 Pfr）。植物光敏色素显示从生理活性 Pfr 到 Pr 的额外热转换途径。在典型的胆红素结合细菌光敏色素中发现了相同的反应模式，与浴光细菌光敏色素的反向热转化相反。然而，原型和浴光细菌光敏色素中 Pfr 状态的不同热稳定性的分子起源尚不清楚。我们使用组合光谱理论方法分析了各种浴光和原型胆红素结合光敏色素的 Pfr 状态的发色团结合口袋的结构。对于假单胞菌的浴光光敏色素的 Pfr 状态，计算和实验胆红素辅因子的拉曼光谱之间非常好的一致性与以前晶体学分析的重要结论一致，特别是发色团的 ZZEssa 构象及其与蛋白质的共价连接方式。高度均匀的发色团构象似乎是浴光光敏色素 Pfr 状态的独特性质。这与原型光敏色素的 Pfr 状态形成鲜明对比，原型光敏色素的 Pfr 状态显示在终端甲桥 A-B 和 C-D 处具有小结构差异的两个亚状态之间的构象平衡。这些差异可能主要源于辅因子与高度保守的天冬氨酸-194 的相互作用，这种相互作用通过其羧基功能在浴光光敏色素中发生。通过原型光敏色素中的羰基功能发生的较弱相互作用可能导致发色团口袋的结构灵活性更高，为热（ZZE→ZZZ）Pfr 到 Pr 的反向转换打开反应通道。
