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Laboratory | Immune system
For a better quality of diffraction
Recently, we have developed a crystallogenesis kit to keep proteins in a good crystalline state. This is the CryoProtX kit. Cryoprotection is the critical step prior to freezing crystals. Currently the kit is sold by the company "Molecular Dimensions" after agreement between CEA and this society.
New developments include the use of antibodies and engineering of bacterial proteins that bind immunoglobulins to promote crystallization of the antigen-antibody complexes, as well as the construction of protein systems scaffolding using fusion proteins and peptide tags to prevent the production of antibodies against each new protein target.
Complexes between proteins, between proteins and nucleic acids or between proteins and peptides are biologically more interesting that the only protein. The complexes can present additional challenges for crystallization, but in the case of macromolecules difficult to crystallize alone, they also bring new opportunities, especially when the system becomes combinatorial with several partners. When two antibodies have distinct epitopes, they can be used in a sandwich-complex.
The improvement of current methods includes the development of screening techniques for a more efficient search of crystallization conditions when you only have small amounts of protein, and a more targeted use of additives which can improve the size and quality of the obtained crystals. This includes additives that control the oxidation or reduction of free cysteines, and those that modify the characteristics of nucleation and crystal growth. The expertise in crystallogenesis is available to collaborators who have structural needs to ensure the success of ongoing projects.
In the field of antibodies, we share with Dr. S Jorieux of the LFB Research Unit a particular interest to the Fc portion of human immunoglobulins G and its mode of attachment to its receptors; this fragment is the target of a pathological response from the immune system, including rheumatoid factors.
The Fc fragment is used as a fusion partner for the expression of the glycoproteins. We have conducted tests to obtain crystals of glycoproteins fused to Fc complexed to the rheumatoid factor.
CEA is a French government-funded technological research organisation in four main areas: low-carbon energies, defense and security, information technologies and health technologies. A prominent player in the European Research Area, it is involved in setting up collaborative projects with many partners around the world.