Tsoraev, G. V. | Search bioRxiv

bioRxiv2018

Radioprotective role of cyanobacterial phycobilisomes

It is now generally accepted that cyanobacteria are responsible for production of oxygen, which led to the so-called \"Great Oxygenation Event\". Appearance of dioxygen in Earths atmosphere resulted in formation of the ozone layer and the ionosphere, which caused significant reduction of ionizing radiation levels at the surface of our planet. This event not only increased biological diversity but also canceled the urgency of previously developed mechanisms of DNA protection, which allowed to survive and develop in harsh environmental conditions including exposure to cosmic rays. In order to test the hypothesis if one of the oldest organisms on Earth retained ancient protection mechanisms, we studied the effect of ionizing radiation (IoR, here: -particles with a kinetic energy of about 30 MeV) and space flight during the mission of the Foton-M4 satellite on cells of Synechocystis sp. PCC6803. By analyzing spectral and functional characteristics of photosynthetic membranes we revealed numerous similarities between cells exposed to IoR and after the space mission. In both cases, we found that excitation energy transfer from phycobilisomes to photosystems was interrupted and the concentration of phycobiliproteins was significantly reduced. Although photosynthetic activity was severely suppressed, the effect was reversible and the cells were able to rapidly recover from stress under normal conditions. Moreover, in vitro experiments demonstrated that the effect of IoR on isolated phycobilisomes was completely different from such in vivo. These observations suggest that the actual existence and the uncoupling of phycobilisomes under irradiation stress could play specific role not only in photo-, but also in radioprotection, which was crucial for early stages of evolution and the development of Life on Earth.

biophysics↗

bioRxiv2017

The photocycle of orange carotenoid protein conceals distinct intermediates and asynchronous changes in the carotenoid and protein components

The 35 kDa water-soluble Orange Carotenoid Protein (OCP) is responsible for photoprotection in cyanobacteria. It acts as a light intensity sensor that simultaneously serves as efficient quencher of phycobilisome excitation energy as well as of reactive oxygen species. Photoactivation triggers large-scale conformational rearrangements to convert OCP from the orange OCPO state to the red active signaling state OCPR, as demonstrated by various structural methods. Eventually, such rearrangements imply complete yet reversible separation of structural domains (C- and N-terminal domain) and significant translocation of the carotenoid cofactor. Very recently, dynamic crystallography of OCPO crystals suggested the existence of photocycle intermediates with small-scale rearrangements that may trigger further transitions in the protein. However, the currently existing gap between the ultra-fast picosecond and 100 millisecond time scale of spectroscopic and structural data precludes knowledge about distinct intermediate states. In this study, we took advantage of single 7 ns laser pulses to study carotenoid absorption transients in OCP on the time-scale from 100 ns to 10 s, which allowed us to detect a red intermediate state preceding the red signaling state OCPR. In addition, time-resolved fluorescence spectroscopy and following assignment of carotenoid-induced quenching of different tryptophan residues revealed a novel orange intermediate state, which appears during back-relaxation of photoactivated OCPR to OCPO. Our results show asynchronous changes in the carotenoid and protein components and provide refined mechanistic information about the OCP photocycle as well as introduce new kinetic signatures for future studies of OCP photoactivity and photoprotection.\n\nSignificance statementCyanobacteria utilize the Orange Carotenoid Protein (OCP) to protect their photosynthetic apparatus from the harmful effects of intense sunlight. OCP is a blue light-triggered photoswitch, which undergoes photoconversion from its dark adapted orange to the active red state, the latter being able to interact with the phycobilisome antennae and quench their fluorescence, thus avoiding excessive energy flow to the photosystems. With the help of the fluorescence recovery protein (FRP), OCP detaches from phycobilisomes and can return faster into the orange state. Until now, only the thermodynamically stable orange state and the metastable red state are established in a primitive photocycle. In this work, we apply transient absorption and fluorescence spectroscopy and identify two novel photocycle intermediates of physiological relevance.

biophysics↗