SARomics Biostructures offers protein crystallization services platform, including protein-ligand complex crystallization and structure determination by X-ray crystallography. Crystallization and crystallography are the oldest methods for three-dimensional structure determination at atomic resolution. After the discovery of X-rays by Conrad Röntgen (Nobel Prize in Physics, 1901), the physical principles of X-ray diffraction were discovered by Max von Laue (Nobel Prize in Physics, 1914), while the foundation for X-ray crystallography was laid by the Braggs father and son (Nobel Prize in Physics, 1915). Apart from X-ray crystallography, currently both NMR spectroscopy and cryo-electron microscopy may provide atomic resolution protein structures. However, the vast majority of entries in the Protein Data Bank (PDB) still originate from X-ray crystallography.
The company’s recombinant protein expression and purification platform (see our catalogue of high purity crystallisation grade proteins) is adapted for obtaining high purity and stable crystallization grade proteins. After cloning, expression and purification, and prior to crystallization, an accurate characterization of the state of the recombinant protein is performed. Probably one of the most important requirements is a monodisperse solution of the protein at a given concentration, which means the solution should not contain aggregated or denatured material. Monodispersity may be assessed using dynamic light scattering (DLS). The protein should also be stable in solution within a certain range of pH and ionic strength. By other words, a proper characterization of the protein, including its solubility, is critical for the success of a crystallization project.
Our services platform is built up around the most advanced biophysical protein characterization methods and includes:
After protein characterization, conditions for obtaining well-diffracting single crystals need to be identified (if the protein has not been crystallized before). The number of experimental parameters affecting protein crystallization can be very large. Among these one can find:
When suitable crystals are obtained it is time for the first test in an X-ray beam. If the X-ray diffraction from the crystals is of good quality (good resolution and nice X-ray diffraction pattern), data are collected at an X-ray source. Among others, we use MAX IV synchrotron radiation facility, which is located in Lund, or send the crystals to other synchrotrons for remote data collect.
If an experimental structure for the same protein is already available, the method of molecular replacement is used, an electron density map is calculated and interpreted, to identify the position of bound ligands, ligand occupancy, etc. Further refinement of the model will ensure that the fine details of the structure are well visible in the electron density map.
Additional information on protein structure and crystallization can be found on our educational site.
Below you can download a guide to shipping samples and an overview of sample handling at our company.
• FastLane™ Premium structures
• FastLane™ Standard Structures
• Antibody-Antigen Complexes
• Biosimilars Quality Validation
• Integrated Lead Discovery
• Fragment Library Screening
• In silico Lead Discovery Services
• NMR Spectroscopy Services
• ProPHECY™ - in silico Peptide and Protein Optimization
96-well protein crystallization plate from Hampton Research for the use of sitting-drop vapor diffusion technique.
For the optimization of the crystallization and structure determination pipeline, SARomics Biostructures has developed standard operation procedures (SOP), which allow an efficient use of time and resources, also helping in keeping the operation costs as low as possible for our clients. These include:
• Developments of Freedom evo scripts and excel scripts for optimization of crystallization experiments
• Standard operating procedures (SOPs) for dynamic light scattering evaluation of protein solutions
• Development of SOPs for X-ray data collection
• Standard protocols for fast track structure determination