Chromophore-Protein Interplay during the Phytochrome Photocycle Revealed by Step-Scan FTIR Spectroscopy

• Phytochrome proteins regulate many photo-responses of plants and microorganisms. Light absorption causes isomerization of the biliverdin chromophore, which triggers a series of structural changes to activate the signaling domains of the protein. However, the structural changes are elusive, and therefore the molecular mechanism of signal transduction remains poorly understood. Here, we apply twocolor step-scan infrared spectroscopy to the bacteriophytochrome from Deinococcus radiodurans. We show by recordings in H2O and D2O that the hydrogen bonds to the biliverdin D-ring carbonyl become disordered in the first intermediate (Lumi-R) forming a dynamic microenvironment, then completely detach in the second intermediate (Meta-R), and finally reform in the signaling state (Pfr). The spectra reveal via isotope labeling that the refolding of the conserved "PHY-tongue" region occurs with the last transition between Meta-R and Pfr. Additional changes in the protein backbone are detected already within microseconds in Lumi-R Aided by molecular dynamics simulations, we find that a strictly conserved salt bridge between an arginine of the PHY tongue and an aspartate of the chromophore binding domains is broken in Lumi-R and the arginine is recruited to the D-ring C=O. This rationalizes how isomerization of the chromophore is linked to the global structural rearrangement in the sensory receptor. Our findings advance the structural understanding of phytochrome photoactivation.

Subject headings

MEDICIN OCH HÄLSOVETENSKAP  -- Medicinska och farmaceutiska grundvetenskaper -- Andra medicinska och farmaceutiska grundvetenskaper (hsv//swe)

MEDICAL AND HEALTH SCIENCES  -- Basic Medicine -- Other Basic Medicine (hsv//eng)

Keyword

time-resolved ftir

photoactive yellow protein

induced proton release

structural-changes

crystal-structure

ir spectroscopy

raman-spectroscopy

nmr-spectroscopy

oat phytochrome

ground-state

Chemistry

Publication and Content Type

ref (subject category)

art (subject category)