High resolution crystallography of macromolecules

The recent developments and availability of modern third-generation synchrotron radiation facilities have a huge impact on macromolecular X-ray crystallography. In connection with a number of methodological improvements and new crystallographic software ranging from data processing to refinement, a unique opportunity has arisen to determine the macromolecular structures with unprecedented high resolution and quality, at a level traditionally reserved for small molecules. At this resolution, individual atoms are clearly resolved and fine details of the structures become visible directly in the electron density maps. The great importance of such structures is the possibility of having broader insights into macromolecule function. At very high resolution, hydrogen atoms can be seen in electron density maps and the detailed information about the protonation states of catalytically important residues can be studied, what often is critical for full understanding of enzymatic reactions. Atomic resolution gives the opportunity for clear definition of multiple conformations, although the proportion of disordered residues is higher at higher resolution, and the disorder is seen as distinct alternative conformations. Water in macromolecular crystal plays an important role in macromolecule’s function and stabilization. Ultrahigh resolution data allows to refine water molecules with anisotropic displacement parameters and refine them with fractional occupancies. In this situation analyzing the subtle hydrogen bond network, involving precisely located water molecules, is possible. Atomic resolution structures can be refined without or with only very weak stereochemical restraints. Macromolecular models refined at ultrahigh resolutions, for well ordered structures, can be used for validation and improvement of stereochemical restraint libraries, commonly used during refinement of macromolecular structures