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Light-dependent flavin redox and adduct states control the conformation and DNA-binding activity of the transcription factor EL222
AS. Chaudhari, A. Favier, ZA. Tehrani, T. Kovaľ, I. Andersson, B. Schneider, J. Dohnálek, J. Černý, B. Brutscher, G. Fuertes
Language English Country England, Great Britain
Document type Journal Article
Grant support
24-11819S
Czech Science Foundation
RVO86652036
Czech Academy of Sciences
LM2023042
MEYS
CZ.02.1.01/0.0/0.0/18_046/0015974
UP CIISB
871037
iNEXT-Discovery
Horizon 2020
FR2054
IR INFRANALYTICS
UAR 3518 CNRS-CEA-UGA-EMBL
Grenoble Instruct-ERIC
ANR-10-INBS-0005-02
FRISBI
ANR-17-EURE-0003
CBH-EUR-GS
90254
e-Infrastruktura CZ
LM2023055
ELIXIR CZ Research Infrastructure
NLK
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PubMed
40119733
DOI
10.1093/nar/gkaf215
Knihovny.cz E-resources
- MeSH
- Bacterial Proteins chemistry metabolism MeSH
- DNA-Binding Proteins chemistry metabolism MeSH
- DNA * chemistry metabolism MeSH
- Flavin Mononucleotide * chemistry metabolism MeSH
- Flavins chemistry metabolism MeSH
- Kinetics MeSH
- Protein Conformation MeSH
- Crystallography, X-Ray MeSH
- Oxidation-Reduction * MeSH
- Molecular Dynamics Simulation MeSH
- Light * MeSH
- Thermosynechococcus metabolism MeSH
- Transcription Factors metabolism chemistry MeSH
- Protein Binding MeSH
- Publication type
- Journal Article MeSH
The activity of the light-oxygen-voltage/helix-turn-helix (LOV-HTH) photoreceptor EL222 is regulated through protein-protein and protein-DNA interactions, both triggered by photo-excitation of its flavin mononucleotide (FMN) cofactor. To gain molecular-level insight into the photocycle of EL222, we applied complementary methods: macromolecular X-ray crystallography (MX), nuclear magnetic resonance (NMR) spectroscopy, optical spectroscopies (infrared and UV-visible), molecular dynamics/metadynamics (MD/metaD) simulations, and protein engineering using noncanonical amino acids. Kinetic experiments provided evidence for two distinct EL222 conformations (lit1 and lit2) that become sequentially populated under illumination. These two lit states were assigned to covalently bound N5 protonated, and noncovalently bound hydroquinone forms of FMN, respectively. Only subtle structural differences were observed between the monomeric forms of all three EL222 species (dark, lit1, and lit2). While the dark state is largely monomeric, both lit states undergo monomer-dimer exchange. Furthermore, molecular modeling revealed differential dynamics and interdomain separation times arising from the three FMN states (oxidized, adduct, and reduced). Unexpectedly, all three EL222 species can associate with DNA, but only upon blue-light irradiation, a high population of stable complexes is obtained. Overall, we propose a model of EL222 activation where photoinduced changes in the FMN moiety shift the population equilibrium toward an open conformation that favors self-association and DNA-binding.
Department of Cell and Molecular Biology Uppsala University Uppsala 75124 Sweden
Faculty of Science Charles University Prague 11636 Czech Republic
Université Grenoble Alpes CEA CNRS Institut de Biologie Structurale Grenoble Cedex 9 38044 France
References provided by Crossref.org
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- $a The activity of the light-oxygen-voltage/helix-turn-helix (LOV-HTH) photoreceptor EL222 is regulated through protein-protein and protein-DNA interactions, both triggered by photo-excitation of its flavin mononucleotide (FMN) cofactor. To gain molecular-level insight into the photocycle of EL222, we applied complementary methods: macromolecular X-ray crystallography (MX), nuclear magnetic resonance (NMR) spectroscopy, optical spectroscopies (infrared and UV-visible), molecular dynamics/metadynamics (MD/metaD) simulations, and protein engineering using noncanonical amino acids. Kinetic experiments provided evidence for two distinct EL222 conformations (lit1 and lit2) that become sequentially populated under illumination. These two lit states were assigned to covalently bound N5 protonated, and noncovalently bound hydroquinone forms of FMN, respectively. Only subtle structural differences were observed between the monomeric forms of all three EL222 species (dark, lit1, and lit2). While the dark state is largely monomeric, both lit states undergo monomer-dimer exchange. Furthermore, molecular modeling revealed differential dynamics and interdomain separation times arising from the three FMN states (oxidized, adduct, and reduced). Unexpectedly, all three EL222 species can associate with DNA, but only upon blue-light irradiation, a high population of stable complexes is obtained. Overall, we propose a model of EL222 activation where photoinduced changes in the FMN moiety shift the population equilibrium toward an open conformation that favors self-association and DNA-binding.
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