Centre de Biologie Structurale

Measuring protein structural dynamics

• On diffusing molecules, in a confocal geometry, to measure distances and fast conformational changes
• On immobilized molecules to measure association / dissociation kinetics and conformational changes in the seconds timescales

All projects are run as collaborations. Please This email address is being protected from spambots. You need JavaScript enabled to view it. for help choosing the most appropriate approach. We also provide advice to design your experiment and support with fluorescent labeling and data analysis. After a meeting, we can run a few experiments to assess the feasibility of the approach. Then a collaboration scheme is defined between the team and the facility. Our R&D division can also implement specific adaptations for your project if needed.

Measurements of distances changes within E. Coli and MTB transcription complexes using single molecule FRET (from Vishwakarma et al.)

Selected publications:

Quast R.B., Fatemi F., Kranendonk M., Margeat E.*, Truan G.* Accurate Determination of CPR Conformational Equilibrium by smFRET using Dual Orthogonal Non-Canonical Amino Acid Labeling, ChemBioChem, 2019, 20(5):659-666

Vishwakarma RK, Cao AM, Morichaud Z, Perumal AS, Margeat E, Brodolin K, Single-molecule analysis reveals the mechanism of transcription activation in M. Tuberculosis. Sci Adv, 2018

Ait-Bara S, Clerte C, Declerck N, Margeat E. Competitive folding of RNA structures at a termination / antitermination site. RNA, 2017

Ait-Bara S, Clerte C, Margeat E. Single-Molecule FRET Characterization of RNA Remodeling Induced by an Antitermination Protein, Methods Mol Biol, 2015

Olofsson L, Felekyan S, Doumazane E, Scholler P, Fabre L, Zwier JM, Rondard P, Seidel CA, Pin JP, Margeat E. Fine tuning of sub-millisecond conformational dynamics controls metabotropic glutamate receptors agonist efficacy, Nat Commun, 2014

SmFRET TCSPC

TIRF / 3D MFM

Measuring molecule diffusions

sptPALM in A.thaliana - credit: Alexandre Martinière

All projects are run as collaborations. Please This email address is being protected from spambots. You need JavaScript enabled to view it. to help you choose the most appropriate approach. We also provide advice for the design of your experiment and support for data analysis. After a meeting, we can conduct some experiments to assess the feasibility of the approach. Then a collaboration scheme is defined between the team and the facility. Our R&D division can also implement specific adaptations for your project if needed.

Selected publications:

B. Guilhas, J.C. Walter, J. Rech, G. David, N.-O. Walliser, J. Palmeri, C., Mathieu-Demaziere, A. Parmeggiani, J.Y. Bouet, A. Le Gall, M. Nollmann. ATP-driven separation of liquid phase condensates in bacteria, Molecular Cell, 2020 Jul 16;79(2):293-303.e4. pubmed Id 32679076

Martiniere A, Fiche JB, Smokvarska M, Mari S, Alcon C, Dumont X, Hematy K, Jaillais Y, Nollmann M, Maurel C Osmotic stress activates two reactive oxygen species pathways with distinct effects on protein nanodomains and diffusion Plant Physiol 2019, 179 ( 4): 1581-1593 pubmed Id 30718348

Platre MP, Bayle V, Armengot L, Bareille J, Marques-Bueno MDM, Creff A, Maneta-Peyret L, Fiche JB, Nollmann M, Miege C, Moreau P, Martiniere A, Jaillais Y Developmental control of plant Rho GTPase nano-organization by the lipid phosphatidylserine Science 2019,364(6435):57-62 pubmed Id 30948546

Faure LM, Fiche JB, Espinosa L, Ducret A, Anantharaman V, Luciano J, Lhospice S, Islam ST, Treguier J, Sotes M, Kuru E, Van Nieuwenhze MS, Brun YV, Theodoly O, L A, Nollmann M, Mignot T The mechanism of force transmission at bacterial focal adhesion complexes Nature 2016, 539(7630):530-535 pubmed Id 27749817

​Antoine Le Gall, Diego I. Cattoni, Baptiste Guilhas, Céline Mathieu-Demazière, Laura Oudjedi, Jean-Bernard Fiche, Jérôme Rech, Sara Abrahamsson, Heath Murray, Jean-Yves Bouet, Marcelo Nollmann. Bacterial partition complexes segregate within the volume of the nucleoid. Nature Communications, 7: 12107, 5 July 2016

Dosset P, Rassam P, Fernandez L, Espenel C, Rubinstein E, Margeat E, Milhiet PE. Automatic detection of diffusion modes within biological membranes using back-propagation neural network, BMC Bioinformatics, 2016

2 photons FCS

SmFRET TCSPC

TIRF / 3D MFM

2 photons STED / FCS

PALM

Measuring biomolecular interactions

1. Local concentrations and oligomerization states (N&B, FCS)
2. Binding between a fluorescent substrate and a partner (Fluorescence Anisotropy, FCS)
3. Binding between 2 fluorescent substrates (FCCS, smFRET, FLIM)
4. Changes in the local environment of fluorescent probes (Fluorescence Anisotropy, Fluorescence lifetime spectroscopy and imaging (FLIM))

All projects are run as collaborations. Please This email address is being protected from spambots. You need JavaScript enabled to view it. to help you choose the most appropriate approach. We also provide advice for the design of your experiment and support for fluorescent labeling and data analysis. After a meeting, we can conduct some experiments to assess the feasibility of the approach. Then a collaboration scheme is defined between the team and the facility. Our R&D division can also implement specific adaptations for your project if needed.

Intensity and brightness (inset) image by two-photon microscopy of a primary neuron transfected with SnapTag-mGlu2 labeled with Alexa488 (from Moller et al.)

Selected publications:

Møller T.C., Hottin J., Clerté C., Zwier J.M., Durroux T., Rondard P., Prezeau L., Royer C.A., Pin J.P., Margeat E., Kniazeff J. Oligomerization of a metabotropic glutamate receptor in neurons controlled by its structural dynamics, Scientifics Reports, 2018

Ferrer M, Clerte C, Chamontin C, Basyuk E, Laine S, Hottin J, Bertrand E, Margeat E, Mougel M, Imaging HIV-1 RNA dimerization in cells by multicolor super-resolution and fluctuation microscopies. Nucleic Acids Res, 2016

2 photons FCS

SmFRET TCSPC

Protein characterization (fluorescence and luminescence)

Berthold Mithras LB940

CLARIOstar

Cytation 3

Synergy H1

TECAN Safire2

Typhoon FLA 9500

Amersham Imager 600