UMBC Chemistry and Biochemistry - Protein Crystallography X-ray Lab

Protein crystallography lies directly at the interface between molecular biology, chemistry, and physics. It is currently responsible for the majority of 3D structures of proteins and nucleic acids solved to date. To solve the crystal structure of a macromolecule by X-ray crystallography, we initially require milligram quantities of highly pure protein. This is achieved using molecular cloning techniques and column chromatography. The purified protein is then placed under conditions that promote a transition from the liquid to the solid phase. These conditions are chosen to promote crystal formation as compared to random aggregation of the protein (referred to as precipitation). Crystals are simply a 3D array of identical molecules that interact with each other in precisely the same way throughout the crystal. Protein crystals vary in size between 0.05 - 0.5 mm. When crystals are placed in a beam of X-rays (wavelength of 0.9 – 2Å), a small portion of the X-ray beam is scattered in a direction different from its original direction. This effect is referred to as diffraction of X-rays. Thousands of diffraction events occur for a protein crystal. The diffraction of X-rays by crystals is mathematically related to two properties of the crystal: how the molecules are packed in the crystal and the structure of the molecules that makes up the crystal. By collecting the diffraction data, we can determine the structure of the molecule that makes up the crystal using computational algorithms.