Methods: rationalisation of protein crystallization

Contact: Monika Spano

A rational way to find the appropriate conditions to grow crystal samples for bio- crystallography is to explore the crystallization phase diagram allowing the precise control of the parameters affecting the crystal growth process. First, the nucleation is induced at supersaturated conditions close to the solubility boundary between the nucleation and metastable regions. Then, crystal growth is further achieved in the metastable zone – which is the optimal location for a slower and ordered crystal expansion – by modulation of specific physical parameters. A thorough knowledge of the phase diagram is vital in any crystallization experiment. The relevance of the selection of the starting position and the kinetic pathway undertaken in controlling most of the final properties of the synthesized crystals has been shown.
The OptiCrys/MicroCrys platforms enable users to benefit from established rational strategies for the optimization of crystal growth using precise in situ control of the temperature and chemical composition of the crystallization solution through dialysis (and microdialysis). Systematic phase diagrams in multidimensional space are studied using small amounts of protein material in the serial approach proposed by the instruments developed and breaking with the current paradigm of parallel experiments.
Established rational crystallization strategies can be beneficial to provide sufficient scattering volumes for neutron protein crystallography that require large-volume well ordered single crystals as well as to generate homogeneous populations of uniformly sized protein crystals required for use by other advanced serial diffraction techniques.

Selected publications:

1. Large crystal growth for neutron protein crystallography. Budayova-Spano M, Koruza K, Fisher Z (2020) Methods Enzymol. 634, 21-46.

2. Optimization of crystallization of biological macromolecules using dialysis combined with temperature control. Junius N, Vahdatahar E, Oksanen E, Ferrer J-L, Budayova-Spano M (2020) J. Appl. Crystallogr. 53, 686-698.

3. Mixing Salts and Poly(ethylene glycol) into Protein Solutions: The Effects of Diffusion across Semipermeable Membranes and of Convection. Apostolopoulou V, Junius N, Sear RP, M. Budayova-Spano (2020) Cryst. Growth Des. 20, 3927-3936.

4. Optimization of crystal growth for neutron macromolecular crystallography. E. Vahdatahar E, Junius N, Budayova-Spano M (2021) J. Vis. Exp. 169, e61685

5. Crystallization of proteins on chip by microdialysis for in situ X-ray diffraction studies. Jaho S, Junius N, Borel F, Sallaz-Damaz Y, Salmon J-B, Budayova-Spano M (2021) J. Vis. Exp. 170, e61660