Activation of tyrosine kinases by mutation of the gatekeeper threonine

Nat Struct Mol Biol. 2008 Oct;15(10):1109-18. doi: 10.1038/nsmb.1486. Epub 2008 Sep 14.


Protein kinases targeted by small-molecule inhibitors develop resistance through mutation of the 'gatekeeper' threonine residue of the active site. Here we show that the gatekeeper mutation in the cellular forms of c-ABL, c-SRC, platelet-derived growth factor receptor-alpha and -beta, and epidermal growth factor receptor activates the kinase and promotes malignant transformation of BaF3 cells. Structural analysis reveals that a network of hydrophobic interactions-the hydrophobic spine-characteristic of the active kinase conformation is stabilized by the gatekeeper substitution. Substitution of glycine for the residues constituting the spine disrupts the hydrophobic connectivity and inactivates the kinase. Furthermore, a small-molecule inhibitor that maximizes complementarity with the dismantled spine (compound 14) inhibits the gatekeeper mutation of BCR-ABL-T315I. These results demonstrate that mutation of the gatekeeper threonine is a common mechanism of activation for tyrosine kinases and provide structural insights to guide the development of next-generation inhibitors.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites
  • CSK Tyrosine-Protein Kinase
  • Cell Line
  • Crystallography, X-Ray
  • Enzyme Activation / genetics
  • ErbB Receptors / chemistry
  • ErbB Receptors / genetics
  • ErbB Receptors / metabolism
  • Humans
  • Hydrophobic and Hydrophilic Interactions
  • Models, Molecular
  • Mutation / genetics*
  • Protein Structure, Tertiary
  • Protein-Tyrosine Kinases / chemistry
  • Protein-Tyrosine Kinases / genetics*
  • Protein-Tyrosine Kinases / metabolism*
  • Proto-Oncogene Proteins c-abl / chemistry
  • Proto-Oncogene Proteins c-abl / genetics
  • Proto-Oncogene Proteins c-abl / metabolism
  • Receptors, Platelet-Derived Growth Factor / chemistry
  • Receptors, Platelet-Derived Growth Factor / genetics
  • Receptors, Platelet-Derived Growth Factor / metabolism
  • Threonine / genetics*
  • Threonine / metabolism*
  • src-Family Kinases


  • Threonine
  • ErbB Receptors
  • Protein-Tyrosine Kinases
  • Receptors, Platelet-Derived Growth Factor
  • CSK Tyrosine-Protein Kinase
  • Proto-Oncogene Proteins c-abl
  • src-Family Kinases
  • CSK protein, human

Associated data

  • PDB/3DQW
  • PDB/3DQX