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Time-resolved SAXS

par Guillaume Tresset - publié le , mis à jour le

 

 

TIME-RESOLVED SAXS


Guillaume Tresset


Small-angle X-ray scattering (SAXS) has become a powerful technique over the last decades for studying biological macromolecules in solution with a nanometer-scaled resolution. The system of interest can be studied in its native conditions and need not be ordered as is the case for X-ray crystallography. Thanks to the development of high brilliance synchrotron sources and to the considerable progress realized on detectors, time-resolved SAXS (TR-SAXS) gives now access to dynamical processes occurring over a few milliseconds.


(Left) Synchrotron SOLEIL near Orsay. (Right) European Synchrotron Radiation Facility (ESRF) in Grenoble.


We develop algorithms that process the large amount of data generated by TR-SAXS experiments. For example, by constructing kinetic models that account for the self-assembly and disassembly of icosahedral viral capsids, we have identified the structure of long-lived intermediate species as well as the evolution of their concentration, consistently with spatio-temporal scattering measurements.


Workflow for analysis and modeling of TR-SAXS data.


Preliminary kinetic data can be obtained in-house with our static light scattering setup coupled to a stopped-flow apparatus. The latter is a SFM-4000 model from Biologic equipped with four syringes and a hard-stop valve. Rapid mixing can be achieved in a few milliseconds. The scattered light intensity is collected at 90° with a photomultiplier tube. Time-resolved fluorescence and absorbance measurements can also be carried out at wavelengths comprised between 300 and 700 nm.


Static light scattering setup coupled to a stopped-flow apparatus.

REFERENCES



M. CHEVREUIL, D. LAW-HINE, J. CHEN, S. BRESSANELLI, S. COMBET, D. CONSTANTIN, J. DEGROUARD, J. MÖLLER, M. ZEGHAL, G. TRESSET (2018) Nonequilibrium self-assembly dynamics of icosahedral viral capsids packaging genome or polyelectrolyte. Nat. Commun. 9 3071.


D. LAW-HINE, M. ZEGHAL, S. BRESSANELLI, D. CONSTANTIN, G. TRESSET (2016) Identification of a major intermediate along the self-assembly pathway of an icosahedral viral capsid by using an analytical model of a spherical patch. Soft Matter 12 6728-6736.


D. LAW-HINE, A. K. SAHOO, V. BAILLEUX, M. ZEGHAL, S. PREVOST, P. K. MAITI, S. BRESSANELLI, D. CONSTANTIN, G. TRESSET (2015) Reconstruction of the disassembly pathway of an icosahedral viral capsid and shape determination of two successive intermediates. J. Phys. Chem. Lett. 6 3471-3476.


G. TRESSET, C. LE COEUR, J.-F. BRYCHE, M. TATOU, M. ZEGHAL, A. CHARPILIENNE, D. PONCET, D. CONSTANTIN, S. BRESSANELLI (2013) Norovirus capsid proteins self-assemble through biphasic kinetics via long-lived stave-like intermediates. J. Am. Chem. Soc. 135 15373-15381.