NMR spectroscopic investigation of mobility and hydrogen bonding of the chromophore in the binding pocket of phytochrome proteins.

For a complete understanding of the light reception of phytochrome proteins, a detailed study of the structure and dynamics of the binding pocket at atomic resolution is required. Structures from X-ray crystallography and NMR spectroscopy are available and have been able to provide a picture of the binding pocket. NMR spectroscopy has, in addition, shown that the chromophore exhibits noticeable dynamics in the binding pocket of the cyanobacterial phytochrome Cph1. Herein, NMR spectroscopy is used to investigate further the mobility of the chromophore by analyzing the line widths of the resonances of the chromophore in various environments, in particular other protein environments. It is shown that the chromophore exhibits a different mobility in the binding pocket of the bacterial phytochrome Agp1 than in that of the cyanobacterial phytochrome Cph1. Finally, it is shown that NMR spectroscopy is capable of detecting hydrogen bonds in the binding pocket of phytochromes by observing slow exchange of protons in the amino acid side chains.