| 1. |
- Claesson, Elin, 1989, et al.
(författare)
-
The primary structural photoresponse of phytochrome proteins captured by a femtosecond X-ray laser
- 2020
-
Ingår i: eLife. - 2050-084X. ; 9
-
Tidskriftsartikel (refereegranskat)abstract
- Phytochrome proteins control the growth, reproduction, and photosynthesis of plants, fungi, and bacteria. Light is detected by a bilin cofactor, but it remains elusive how this leads to activation of the protein through structural changes. We present serial femtosecond X-ray crystallographic data of the chromophore-binding domains of a bacterial phytochrome at delay times of 1 ps and 10 ps after photoexcitation. The data reveal a twist of the D-ring, which leads to partial detachment of the chromophore from the protein. Unexpectedly, the conserved so-called pyrrole water is photodissociated from the chromophore, concomitant with movement of the A-ring and a key signaling aspartate. The changes are wired together by ultrafast backbone and water movements around the chromophore, channeling them into signal transduction towards the output domains. We suggest that the observed collective changes are important for the phytochrome photoresponse, explaining the earliest steps of how plants, fungi and bacteria sense red light.
|
|
| 2. |
- Cellini, Andrea, 1991, et al.
(författare)
-
Directed ultrafast conformational changes accompany electron transfer in a photolyase as resolved by serial crystallography.
- 2024
-
Ingår i: Nature chemistry. - : Springer Nature. - 1755-4349 .- 1755-4330.
-
Tidskriftsartikel (refereegranskat)abstract
- Charge-transfer reactions in proteins are important for life, such as in photolyases which repair DNA, but the role of structural dynamics remains unclear. Here, using femtosecond X-ray crystallography, we report the structural changes that take place while electrons transfer along a chain of four conserved tryptophans in the Drosophila melanogaster (6-4) photolyase. At femto- and picosecond delays, photoreduction of the flavin by the first tryptophan causes directed structural responses at a key asparagine, at a conserved salt bridge, and by rearrangements of nearby water molecules. We detect charge-induced structural changes close to the second tryptophan from 1 ps to 20 ps, identifying a nearby methionine as an active participant in the redox chain, and from 20 ps around the fourth tryptophan. The photolyase undergoes highly directed and carefully timed adaptations of its structure. This questions the validity of the linear solvent response approximation in Marcus theory and indicates that evolution has optimized fast protein fluctuations for optimal charge transfer.
|
|
| 3. |
- Cellini, Andrea, 1991, et al.
(författare)
-
Structural basis of the radical pair state in photolyases and cryptochromes
- 2022
-
Ingår i: Chemical Communications. - : Royal Society of Chemistry (RSC). - 1359-7345 .- 1364-548X. ; , s. 4889-4892
-
Tidskriftsartikel (refereegranskat)abstract
- We present the structure of a photoactivated animal (6-4) photolyase in its radical pair state, captured by serial crystallography. We observe how a conserved asparigine moves towards the semiquinone FAD chromophore and stabilizes it by hydrogen bonding. Several amino acids around the final tryptophan radical rearrange, opening it up to the solvent. The structure explains how the protein environment stabilizes the radical pair state, which is crucial for function of (6-4) photolyases and cryptochromes.
|
|
| 4. |
- Cellini, Andrea, 1991
(författare)
-
Structural characterization of electron transfer in D.m (6-4) photolyase by time-resolved X-ray crystallography
- 2022
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Photolyases are flavoproteins widely spread in bacteria, archea, fungi, plants and animals. These enzymes absorb blue light and use it as an energy source for repairing damages that are induced on DNA after a prolonged exposure to UV-light. The mechanism of DNA repair in photolyases requires a first step of photo-excitation known as photoactivation. During this process, the chromophore uptakes an electron from a close tryptophan. This event triggers an electron transfer along a chain of tryptophans and results into the reduction of the chromophore to its catalytic form. This thesis is focusing on unravelling the structural changes associated with photoactivation in a photolyase from Drosophila Melanogaster. We employed time-resolved serial crystallography as the main technique of investigation. In the three papers, we present the crystallography techniques and conditions that were used for time-resolved experiments in synchrotron and in x-ray free electron laser facilities (XFEL). At first, we solved the structure of the protein in its resting state and then we characterized the structural changes that occur after light activation. We recorded data at different time delays from illumination ranging from ps to ms. The findings show structural changes around the chromophore and the tryptophans involved in the electron transfer. These results contribute to the understanding of the structural adaptation of photolyase during the first electron transfer process. However,further studies are needed to structurally characterized the second step of photoactivation and the process of DNA repair.
|
|
| 5. |
- Cellini, Andrea, 1991, et al.
(författare)
-
The three-dimensional structure of Drosophila melanogaster (6-4) photolyase at room temperature
- 2021
-
Ingår i: Acta Crystallographica Section D-Structural Biology. - : International Union of Crystallography (IUCr). - 2059-7983. ; 77, s. 1001-1009
-
Tidskriftsartikel (refereegranskat)abstract
- (6-4) photolyases are flavoproteins that belong to the photolyase/cryptochrome family. Their function is to repair DNA lesions using visible light. Here, crystal structures of Drosophila melanogaster (6-4) photolyase [Dm(6-4)photolyase] at room and cryogenic temperatures are reported. The room-temperature structure was solved to 2.27 angstrom resolution and was obtained by serial femtosecond crystallography (SFX) using an X-ray free-electron laser. The crystallization and preparation conditions are also reported. The cryogenic structure was solved to 1.79 angstrom resolution using conventional X-ray crystallography. The structures agree with each other, indicating that the structural information obtained from crystallography at cryogenic temperature also applies at room temperature. Furthermore, UV-Vis absorption spectroscopy confirms that Dm(6-4)photolyase is photoactive in the crystals, giving a green light to time-resolved SFX studies on the protein, which can reveal the structural mechanism of the photoactivated protein in DNA repair.
|
|
| 6. |
- Henry, Léocadie, et al.
(författare)
-
New Light on the Mechanism of Phototransduction in Phototropin
- 2020
-
Ingår i: Biochemistry. - : American Chemical Society (ACS). - 0006-2960 .- 1520-4995. ; 59:35, s. 3206-3215
-
Tidskriftsartikel (refereegranskat)abstract
- Phototropins are photoreceptor proteins that regulate blue light-dependent biological processes for efficient photosynthesis in plants and algae. The proteins consist of a photosensory domain that responds to the ambient light and an output module that triggers cellular responses. The photosensory domain of phototropin from Chlamydomonas reinhardtii contains two conserved LOV (light-oxygen-voltage) domains with flavin chromophores. Blue light triggers the formation of a covalent cysteine-flavin adduct and upregulates the phototropin kinase activity. Little is known about the structural mechanism that leads to kinase activation and how the two LOV domains contribute to this. Here, we investigate the role of the LOV1 domain from C. reinhardtii phototropin by characterizing the structural changes occurring after blue light illumination with nano- to millisecond time-resolved X-ray solution scattering. By structurally fitting the data with atomic models generated by molecular dynamics simulations, we find that adduct formation induces a rearrangement of the hydrogen bond network from the buried chromophore to the protein surface. In particular, the change in conformation and the associated hydrogen bonding of the conserved glutamine 120 induce a global movement of the beta-sheet, ultimately driving a change in the electrostatic potential on the protein surface. On the basis of the change in the electrostatics, we propose a structural model of how LOV1 and LOV2 domains interact and regulate the full-length phototropin from C. reinhardtii. This provides a rationale for how LOV photosensor proteins function and contributes to the optimal design of optogenetic tools based on LOV domains.
|
|
