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

J Biol Chem. 2007 Feb 9;282(6):4085-93. doi: 10.1074/jbc.M608182200. Epub 2006 Nov 28.


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.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cells, Cultured
  • Crystallization
  • Cytoplasm / chemistry
  • Cytoplasm / genetics
  • Humans
  • Molecular Sequence Data
  • Mutant Chimeric Proteins / chemical synthesis
  • Mutant Chimeric Proteins / genetics
  • Protein Engineering*
  • Protein Kinase Inhibitors / chemistry
  • Protein Structure, Tertiary / genetics
  • Receptor Protein-Tyrosine Kinases / antagonists & inhibitors
  • Receptor Protein-Tyrosine Kinases / chemical synthesis*
  • Receptor Protein-Tyrosine Kinases / genetics*
  • Receptor, Macrophage Colony-Stimulating Factor / antagonists & inhibitors
  • Receptor, Macrophage Colony-Stimulating Factor / chemistry*
  • Receptor, Macrophage Colony-Stimulating Factor / genetics*
  • Sequence Alignment
  • Spodoptera


  • Mutant Chimeric Proteins
  • Protein Kinase Inhibitors
  • Receptor Protein-Tyrosine Kinases
  • Receptor, Macrophage Colony-Stimulating Factor