Proline: Mother Nature's cryoprotectant applied to protein crystallography.
Publication Type:Journal Article
Source:Acta Crystallogr D Biol Crystallogr, Volume 68, Issue Pt 8, p.1010-8 (2012)
Keywords:1-Pyrroline-5-Carboxylate Dehydrogenase, Acid Phosphatase, Aldose-Ketose Isomerases, Animals, Chickens, Cryoprotective Agents, Crystallization, Crystallography, X-Ray, Egg White, Histidine, Humans, Hydrogen-Ion Concentration, Muramidase, Proline, Protein Binding, Proteins, Temperature
<p>L-Proline is one of Mother Nature's cryoprotectants. Plants and yeast accumulate proline under freeze-induced stress and the use of proline in the cryopreservation of biological samples is well established. Here, it is shown that L-proline is also a useful cryoprotectant for protein crystallography. Proline was used to prepare crystals of lysozyme, xylose isomerase, histidine acid phosphatase and 1-pyrroline-5-carboxylate dehydrogenase for low-temperature data collection. The crystallization solutions in these test cases included the commonly used precipitants ammonium sulfate, sodium chloride and polyethylene glycol and spanned the pH range 4.6-8.5. Thus, proline is compatible with typical protein-crystallization formulations. The proline concentration needed for cryoprotection of these crystals is in the range 2.0-3.0 M. Complete data sets were collected from the proline-protected crystals. Proline performed as well as traditional cryoprotectants based on the diffraction resolution and data-quality statistics. The structures were refined to assess the binding of proline to these proteins. As observed with traditional cryoprotectants such as glycerol and ethylene glycol, the electron-density maps clearly showed the presence of proline molecules bound to the protein. In two cases, histidine acid phosphatase and 1-pyrroline-5-carboxylate dehydrogenase, proline binds in the active site. It is concluded that L-proline is an effective cryoprotectant for protein crystallography.</p>