Novel protein crystal growth technology: Proof of concept

A technology for crystal growth, which overcomes certain shortcomings of other techniques, is developed and its applicability to proteins is examined. There were several unknowns to be determined: the design of the apparatus for suspension of crystals of varying (growing) diameter, control of the temperature and supersaturation, the methods for seeding and/or controlling nucleation, the effect on protein solutions of the temperature oscillations arising from the circulation, and the effect of the fluid shear on the suspended crystals. Extensive effort was put forth to grow lysozyme crystals. Under conditions favorable to the growth of tetragonal lysozyme, spontaneous nucleation could be produced but the number of nuclei could not be controlled. Seed transfer techniques were developed and implemented. When conditions for the orthorhombic form were tried, a single crystal 1.5 x 0.5 x 0.2 mm was grown (after in situ nucleation) and successfully extracted. A mathematical model was developed to predict the flow velocity as a function of the geometry and the operating temperatures. The model can also be used to scaleup the apparatus for growing larger crystals of other materials such as water soluble non-linear optical materials. This crystal suspension technology also shows promise for high quality solution growth of optical materials such as TGS and KDP.