Team 2: Advanced solution NMR methods

Team members

Arijit Maity (Postdoctoal researcher)
Isabel Ayala (molecular biochemistry engineer)
Bernhard Brutscher (CEA research director)
Adrien Favier (research engineer)
Alicia Vallet (NMR platform engineer)

Presentation

The main research focus of our team is the development of optimized solution (and to a minor extent solid-state) NMR pulse sequences and spectral analysis tools for biomolecular applications. Most of these spectroscopic tools are made available to the academic community via our NMRlib package.
Past research efforts have focused on the development of fast multidimensional data acquisition techniques, especially polarization-enhanced fast-pulsing methods (SOFAST, BEST, …). Such methods not only prove useful for speeding up NMR data collection, but they also provide new opportunities for site-resolved kinetic real-time NMR investigations of biomolecular rreactions. Kinetic processes can be initiated in the NMR sample tube for example by a pH or temperature jump, rapid mixing of interacting molecules, or by light. Therefore, we have assembled dedicated hardware for in-situ (inside the NMR magnet) fast mixing of solutions and sample illumination by laser light. As a logical consequence of these NMR developments, we became interested in the mechanistic understanding of light-induced transformations of fluorescent and other light-sensing proteins.
A significant part of the group’s activity is devoted to collaborative research projects in partnership with external users of the NMR facility.

Current projects

NMR investigation of photoswitching mechanisms in light-sensitive proteins.
Funding: ANR20 "StableFP" (2020-2025); ANR22 "Photoswitch NMR" (2023-2027); PEPR-LUMA
Light is a major energy source for living organisms. Light-sensitive proteins are found in all kingdoms of life where they perform various biological functions, such as light harvesting for photosynthesis, vision or circadian clock regulation, but also regulation of gene expression and other signaling pathways. In addition, they provide biotechnological tools for cellular imaging, biosensing, and optogenetic control. We have recently set up a laser-based device that allows in-situ NMR sample illumination at 3 different wave lengths (405, 481, and 561 nm). This device will soon be upgraded to 6 light sources with higher emission power and covering a wide spectral range (from infra-red to UV), as well as additional capabilities for light detection, and a gas-pressure device to probe differences in dioxygen (O2) accessibility in proteins. We will exploit this NMR setup to gain new insights into the photophysical and photochemical transformations that chromophore-containing biomolecules undergo upon exposure to light, by characterizing the conformational dynamics in different photo-stationary states, the interconversion kinetics and pathways, as well as identifying additional “dark” states that are invisible to optical spectroscopy.
Collaborations: D. Bourgeois (IBS),L. Jullien (ENS Paris), G. Fuertes (Prague, CZ)

Recent publications

PRESERVE: Adding variable flip-angle excitation to TROSY NMR spectroscopy.
Brutscher B, Magn. Reson. 2024, doi:10.5194/mr-2024-9.

MAS NMR experiments of corynebacterial cell walls: Complementary 1H- and CPMAS CryoProbe-enhanced 13C-detected experiments.
Vallet A, Ayala I, Perrone B, Hassan A, Simorre JP, Bougault C, Schanda P. J Magn Reson. 2024 Jul;364:107708. doi: 10.1016/j.jmr.2024.107708.

Structure shows that the BIR2 domain of E3 ligase XIAP binds across the RIPK2 kinase dimer interface.
Lethier M, Huard K, Hons M, Favier A, Brutscher B, Erba EB, Abbott DW, Cusack S, Pellegrini, E. Life Sci. Alliance 2023, 6, 1–14. doi: 10.26508/lsa.202201784.

Structural heterogeneity in a phototransformable fluorescent protein impacts its photochemical properties.
Maity A, Wulffelé J, Ayala I, Favier A, Adam V, Bourgeois D, Brutscher B. Advanced Science. 2023/12; e2306272-e2306272. doi: 10.1002/advs.202306272.

The plasma membrane-associated cation-binding protein PCaP1 of Arabidopsis thaliana is a uranyl-binding protein.
Vallet A, Martin-Laffon J, Favier A, Revel B, Bonnot T, Vidaud C, Armengaud C, Gaillard JC, Delangle P, Devime F, Brutscher B et al. J. Hazard. Mater. 2023, 446, doi: 10.1016/j.jhazmat.2022.130668.