Spectroscopic Investigation on the Primary Photoreaction of Bathy Phytochrome Agp2-Pr of Agrobacterium fabrum: Isomerization in a pH-dependent H-bond Network

Bathy phytochrome Agp2 from Agrobacterium fabrum exhibits an unusually low pKa=7.6 in the Pr state in contrast to a pKa>11 in the Pfr state, indicating a pH‐dependent charge distribution and H‐bond network in the Pr chromophore binding pocket around neutral pH. Here, we report on ultrafast UV/Vis absorption spectroscopy of the primary Pr photoisomerization of Agp2 at pH 6 and pH 9 and upon H2O/D2O buffer exchange. The triexponential Pr kinetics slows down at increased pH and pronounced pH‐dependent kinetic isotope effects are observed. The results on the Pr photoreaction suggest: 1) component‐wise hindered dynamics on the chromophore excited‐state potential energy surface at high pH and 2) proton translocation processes either via single‐proton transfer or via significant reorganization of H‐bond networks. Both effects reflect the interplay between the pH‐dependent charge distribution in the Pr chromophore binding pocket on the one hand and chromophore excitation and its Z→E isomerization on the other hand. Networking is important: The unusually low pKa=7.6 of the Pr form of bathy phytochrome Agp2 of Agrobacterium fabrum causes the excited‐state kinetics at high pH to slow down significantly as well as pH‐dependent kinetic isotope effects. Both observations demonstrate the strong contributions of H‐bond network rearrangements and proton transfer to the multidimensional isomerization reaction coordinate.