Structural characterization of a novel Cbl phosphotyrosine recognition motif in the APS family of adapter proteins

J Biol Chem. 2005 May 13;280(19):18943-9. doi: 10.1074/jbc.M414157200. Epub 2005 Feb 28.


The Cbl adapter proteins typically function to down-regulate activated protein tyrosine kinases and other signaling proteins by coupling them to the ubiquitination machinery for degradation by the proteasome. Cbl proteins bind to specific tyrosine-phosphorylated sequences in target proteins via the tyrosine kinase-binding (TKB) domain, which comprises a four-helix bundle, an EF-hand calcium-binding domain, and a non-conventional Src homology-2 domain. The previously derived consensus sequence for phosphotyrosine recognition by the Cbl TKB domain is NXpY(S/T)XXP (X denotes lesser residue preference), wherein specificity is conferred primarily by residues C-terminal to the phosphotyrosine. Cbl is recruited to and phosphorylated by the insulin receptor in adipose cells through the adapter protein APS. APS is phosphorylated by the insulin receptor on a C-terminal tyrosine residue, which then serves as a binding site for the Cbl TKB domain. Using x-ray crystallography, site-directed mutagenesis, and calorimetric studies, we have characterized the interaction between the Cbl TKB domain and the Cbl recruitment site in APS, which contains a sequence motif, RA(V/I)XNQpY(S/T), that is conserved in the related adapter proteins SH2-B and Lnk. These studies reveal a novel mode of phosphopeptide interaction with the Cbl TKB domain, in which N-terminal residues distal to the phosphotyrosine directly contact residues of the four-helix bundle of the TKB domain.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing
  • Adaptor Proteins, Vesicular Transport / chemistry*
  • Adaptor Proteins, Vesicular Transport / physiology
  • Adipose Tissue / cytology
  • Amino Acid Motifs
  • Amino Acid Sequence
  • Animals
  • Binding Sites
  • CHO Cells
  • Calcium / chemistry
  • Calorimetry
  • Cricetinae
  • Crystallography, X-Ray
  • Electrons
  • Immunoblotting
  • Immunoprecipitation
  • Kinetics
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Phosphorylation
  • Phosphotyrosine / chemistry*
  • Protein Binding
  • Protein Conformation
  • Protein Structure, Tertiary
  • Signal Transduction
  • Thermodynamics
  • Transfection
  • Tyrosine / chemistry


  • Adaptor Proteins, Signal Transducing
  • Adaptor Proteins, Vesicular Transport
  • SH2B2 protein, human
  • Phosphotyrosine
  • Tyrosine
  • Calcium

Associated data

  • PDB/1YVH