M1207 - Protein Crystallization

Zhang Fan and Tilman Lamparter

The tertiary structure of a protein can only be obtained by NMR and X-ray crystallography. The protein database contains presently 82000 crystal structures and 10000 NMR structures. One bottleneck for structural studies is the crystallization of a protein which is successful in only 30%. Most often, proteins for crystallization are expressed in E. coli or in eukaryotic host cells and purified by affinity chromatography. In this practical course we will perform purification and protein crystallization with photolyases and phytochromes. You will follow crystal growth in the microscope and learn optimization screening for cases where crystallization has as yet not been established. You will also perform site directed mutagenesis of expression vectors. This technique can be used for optimizations of crystal growth and for analyzing the role of a specific amino acid. We will visualize 3D structures with Pymol and other programs.