Monitoring the Production of High Diffraction-Quality Crystals of Two Enzymes in Real Time Using In Situ Dynamic Light Scattering
by
Raphaël de Wijn 1,†
, Kévin Rollet 1,2,†, Sylvain Engilberge 3, Alastair G. McEwen 4, Oliver Hennig 2, Heike Betat 2, Mario Mörl 2, François Riobé 5, Olivier Maury 5, Eric Girard 3, Philippe Bénas 1, Bernard Lorber 1 and Claude Sauter 1,*
Raphaël de Wijn 1,†, Kévin Rollet 1,2,†, Sylvain Engilberge 3, Alastair G. McEwen 4, Oliver Hennig 2, Heike Betat 2, Mario Mörl 2, François Riobé 5, Olivier Maury 5, Eric Girard 3, Philippe Bénas 1, Bernard Lorber 1 and Claude Sauter 1,*
1
Institut de Biologie Moléculaire et Cellulaire, CNRS, UPR 9002, Architecture et Réactivité de l’ARN, Université de Strasbourg, F-67084 Strasbourg, France
2
Institute for Biochemistry, Leipzig University, D-04103 Leipzig, German
3
Institut de Biologie Structurale, CNRS, CEA, Université Grenoble Alpes, F-38044 Grenoble, France
4
Plateforme de Biologie et Génomique Structurale, Institut de Génétique et de Biologie Moléculaire et Cellulaire, Université de Strasbourg, CNRS UMR 7104, INSERM U 1258, F-67404 Illkirch, France
5
Univ Lyon, ENS Lyon, CNRS-UMR 5182, Université Lyon 1, F-69342 Lyon, France
*
Author to whom correspondence should be addressed.
†
These authors contributed equally to the work.
Crystals 2020, 10(2), 65; https://doi.org/10.3390/cryst10020065
Received: 25 December 2019 / Revised: 13 January 2020 / Accepted: 14 January 2020 / Published: 23 January 2020
(This article belongs to the Special Issue Novel Strategies for Improved Protein Crystallization)
The reproducible preparation of well-diffracting crystals is a prerequisite for every structural study based on crystallography. An instrument called XtalController has recently been designed that allows the monitoring of crystallization assays using dynamic light scattering and microscopy, and integrates piezo pumps to alter the composition of the mother liquor during the experiment. We have applied this technology to study the crystallization of two enzymes, the CCA-adding enzyme of the psychrophilic bacterium Planococcus halocryophilus, and the lysozyme from hen egg white in the presence of a synthetic chemical nucleant. We were able to (i) detect early nucleation events and (ii) drive the crystallization system (through cycles of dissolution/crystallization) toward growth conditions yielding crystals with excellent diffraction properties. This technology opens a way to the rational production of samples for crystallography, ranging from nanocrystals for electron diffraction, microcrystals for serial or conventional X-ray diffraction, to larger crystals for neutron diffraction.
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Keywords:
enzyme; crystallization; dynamic light scattering; nucleation; nucleant; Tb-Xo4 crystallophore; microcrystals; nanocrystals; X-ray diffraction; XtalController
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MDPI and ACS Style
de Wijn, R.; Rollet, K.; Engilberge, S.; McEwen, A.G.; Hennig, O.; Betat, H.; Mörl, M.; Riobé, F.; Maury, O.; Girard, E.; Bénas, P.; Lorber, B.; Sauter, C. Monitoring the Production of High Diffraction-Quality Crystals of Two Enzymes in Real Time Using In Situ Dynamic Light Scattering. Crystals 2020, 10, 65.
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