Abstract
Photosynthesis requires a balance between efficient light harvesting and protection against photodamage. The cyanobacterial photoprotection system uniquely relies on the functioning of the photoactive orange carotenoid protein (OCP) that under intense illumination provides fluorescence quenching of the light-harvesting antenna complexes, phycobilisomes. The recently identified fluorescence recovery protein (FRP) binds to the photoactivated OCP and accelerates its relaxation into the basal form, completing the regulatory circle. The molecular mechanism of FRP functioning is largely controversial. Moreover, since the available knowledge has mainly been gained from studying Synechocystis proteins, the cross-species conservation of the FRP mechanism remains unexplored. Besides phylogenetic analysis, we performed a detailed structural-functional analysis of two selected low-homology FRPs by comparing them with Synechocystis FRP (SynFRP). While adopting similar dimeric conformations in solution and preserving binding preferences of SynFRP toward various OCP variants, the low-homology FRPs demonstrated distinct binding stoichiometries and differentially accentuated features of this functional interaction. By providing clues to understand the FRP mechanism universally, our results also establish foundations for upcoming structural investigations necessary to elucidate the FRP-dependent regulatory mechanism.
Footnotes
List of abbreviations:
- OCP
- carotenoid protein, holoprotein
- OCPAA
- OCP with amino acid substitutions Y201A and W288A, holoprotein
- NTE
- N-terminal extension (comprising the αA helix up to amino acid 20)
- ∆NTE
- OCP with the 12 most N-terminal amino acids deleted, holoprotein
- CTD
- C-terminal domain
- NTD
- N-terminal domain
- HCP
- helical carotenoid protein, NTD homologue of OCP
- COCP
- CTD of Synechocystis OCP (amino acids 165–317), holoprotein
- RCP
- red carotenoid protein, holoprotein
- RCP(apo)
- NTD of Synechocystis OCP (amino acids 1–164), apoprotein
- FRP
- fluorescence recovery protein
- SynFRP
- Synechocystis FRP
- AnaFRP
- Anabaena FRP
- AmaxFRP
- Arthrospira FRP
- Arthrospira
- Arthrospira maxima CS-328
- Anabaena
- Anabaena variabilis PCC 7937
- Synechocystis
- Synechocystis sp. PCC 6803
- CAN
- canthaxanthin
- ECN
- echinenone
- hECN
- 3’-hydroxyechinenone
- AL
- actinic light
- LED
- light-emitting diode
- DLS
- dynamic light scattering
- DTT
- dithiothreitol
- PB
- phycobilisome
- QELS
- quasi-elastic light scattering
- RC
- reaction center of photosystem
- ROS
- reactive oxygen species
- SEC
- size-exclusion chromatography
- SEC-MALLS
- SEC with multiangle laser light scattering analysis
- SAXS
- small-angle X-ray scattering
- SDS-PAGE
- sodium dodecyl sulfate polyacrylamide gel electrophoresis
- MSA
- multiple sequence alignment
- UV
- ultraviolet