Monitoring the Production of High Diffraction-Quality Crystals of Two Enzymes in Real Time Using In Situ Dynamic Light Scattering

Abstract

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.