Practical Use of Glycerol in Protein Crystallization

Glycerol is often used to protect proteins while stored frozen, to enhance their solubility and as a cryosolvent in cryocrystallography, but less so in protein crystallization, probably because it presents problems during screening. Understanding how glycerol affects protein crystallization is important for manual and for high throughput operations, and it will permit greater control of equilibration by vapor diffusion. Certain synthetic inhibitors of matrix metalloproteinase 12 (MMP-12) on mixing with the enzyme cause precipitation and induce nucleation, resulting in unusable microcrystal showers. Glycerol has been used to reduce this effect. We detail the practical problems and advantages in the use of glycerol with MMP-12 and how these observations may be extended to a larger range of problems, focusing on its use by the vapor diffusion method. Glycerol appears to be more effective when coupled with alternative and complementary methods that can similarly reduce excessive nucleation.