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J Biol Chem. 2007 Feb 9;282(6):4085-93. Epub 2006 Nov 28.

Protein engineering of the colony-stimulating factor-1 receptor kinase domain for structural studies.

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Structural Biology, Johnson & Johnson Pharmaceuticals Research and Development, LLC, Exton, Pennsylvania 19341, USA.


A parallel approach to designing crystallization constructs for the c-FMS kinase domain was implemented, resulting in proteins suitable for structural studies. Sequence alignment and limited proteolysis were used to identify and eliminate unstructured and surface-exposed domains. A small library of chimeras was prepared in which the kinase insert domain of FMS was replaced with the kinase insert domain of previously crystallized receptor-tyrosine kinases. Characterization of the newly generated FMS constructs by enzymology and thermoshift assays demonstrated similar activities and compound binding to the FMS full-length cytoplasmic domain. Two chimeras were evaluated for crystallization in the presence and absence of a variety of ligands resulting in crystal structures, and leading to a successful structure-based drug design project for this important inflammation target.

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