Precipitant-Free Lysozyme Crystals Grown by Centrifugal Concentration Reveal Structural Changes

The three-dimensional (3D) structure of a protein molecule in its crystal need not correspond to that found in vivo in many cases, since we usually crystallize protein molecules using precipitants (salts, organic solvents, polymeric electrolytes, etc.), and the precipitants are often incorporated into crystals along with the protein molecules. Although precipitant-free crystallization methods would solve these problems, such methods had not yet been established. We have achieved a novel precipitant-free crystallization method by liquid–liquid phase separation during the centrifugal concentration of lysozyme in ultrapure water. In the 3D structure of the precipitant-free crystal, lysozyme loses a sodium cation and changes the position of Ser72. Deionization of the solution also appears to induce a change in the position of Asp101 and an increase in the activity of lysozyme.