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Unusual spectral properties of bacteriophytochrome Agp2 result from a deprotonation of the chromophore in the red-absorbing form Pr.
Zitatschlüssel Zienicke2013
Autor Zienicke, Benjamin and Molina, Isabel and Glenz, Rene and Singer, Patrick and Ehmer, Dorothee and Escobar, Francisco Velazquez and Hildebrandt, Peter and Diller, Rolf and Lamparter, Tilman
Seiten 31738–51
Jahr 2013
DOI 10.1074/jbc.M113.479535
Journal The Journal of biological chemistry
Jahrgang 288
Nummer 44
Zusammenfassung Phytochromes are widely distributed photoreceptors with a bilin chromophore that undergo a typical reversible photoconversion between the two spectrally different forms, Pr and Pfr. The phytochrome Agp2 from Agrobacterium tumefaciens belongs to the group of bathy phytochromes that have a Pfr ground state as a result of the Pr to Pfr dark conversion. Agp2 has untypical spectral properties in the Pr form reminiscent of a deprotonated chromophore as confirmed by resonance Raman spectroscopy. UV/visible absorption spectroscopy showed that the pKa is >11 in the Pfr form and 7.6 in the Pr form. Unlike other phytochromes, photoconversion thus results in a pKa shift of more than 3 units. The Pr/Pfr ratio after saturating irradiation with monochromatic light is strongly pH-dependent. This is partially due to a back-reaction of the deprotonated Pr chromophore at pH 9 after photoexcitation as found by flash photolysis. The chromophore protonation and dark conversion were affected by domain swapping and site-directed mutagenesis. A replacement of the PAS or GAF domain by the respective domain of the prototypical phytochrome Agp1 resulted in a protonated Pr chromophore; the GAF domain replacement afforded an inversion of the dark conversion. A reversion was also obtained with the triple mutant N12S/Q190L/H248Q, whereas each single point mutant is characterized by decelerated Pr to Pfr dark conversion.
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