Rationalizing Protein Crystallization Screenings through Water Equilibration Theory and Protein Solubility Data

摘要

This work combined water equilibration fundamentals of vapor diffusion crystallization techniques with protein solubility data in order to obtain the variation of protein supersaturation throughout the protein crystallization assays. Once the supersaturation build up profiles (SBUPs) are known, the wide screening space of crystallization conditions is reduced to the key variable for crystal formation and growth, which is supersaturation and its variation with time. Our previous water equilibration model was expanded to include the case of drop evaporation at constant contact area during the hanging drop method. Crystallization experiments of lysozyme were performed under different experimental conditions and the results were interpreted according to the respective SBPUs. In particular, the number and size of the crystals were evaluated at the moment of the SBUP that corresponded to their formation. Following this methodology, two nucleation behaviors were identified depending on the supersaturation levels at which crystal formation occurs. These behaviors, which are believed to be closely linked with the diffracting properties of the crystals, are dictated not only by classic thermodynamic and kinetic factors affecting crystallization and water equilibration, but also by phenomena related to the drop preparation procedures.