X-ray biocrystallography is maybe the most powerful method to obtain a macromolecular structure. The improvement of computational technologies and the development of powerful computer programs together with the enormous increase in the number of protein structures deposited in the Protein Data Bank, render structure determination of new structures much easier than in the past.
Single crystals obtained from the crystallization experiments can be mounted in nylon or plastic microloops in order to be vitrified in suitable cryoprotecting solutions with liquid nitrogen. The crystals are afterwards shipped in a dry-shipper to a synchrotron facility (usually Diamond Light Source, Oxford, UK, or EMBL-Hamburg outstation, Germany) for crystallographic data collection to occur. Diffraction images are processed with the appropriate software and the unique dataset can be used for protein structure solution. The full sequence of the expressed construct is required.
Protein structure refinement and structure analysis complete the procedure.
Protein structure determination is accomplished with the use of Coot, CCP4, iMosflm and/or Phenix computer softwares.