Simultaneous binding of two peptidyl ligands by a SRC homology 2 domain

Biochemistry. 2011 Sep 6;50(35):7637-46. doi: 10.1021/bi200439v. Epub 2011 Aug 9.

Abstract

Src homology 2 (SH2) domains mediate protein-protein interactions by recognizing phosphotyrosine (pY)-containing sequences of target proteins. In all of the SH2 domain-pY peptide interactions described to date, the SH2 domain binds to a single pY peptide. Here, determination of the cocrystal structure of the N-terminal SH2 domain of phosphatase SHP-2 bound to a class IV peptide (VIpYFVP) revealed a noncanonical 1:2 (protein-peptide) complex. The first peptide binds in a canonical manner with its pY side chain inserted in the usual binding pocket, while the second pairs up with the first to form two antiparallel β-strands that extend the central β-sheet of the SH2 domain. This unprecedented binding mode was confirmed in the solution phase by NMR experiments and shown to be adopted by pY peptides derived from cellular proteins. Site-directed mutagenesis and surface plasmon resonance studies revealed that the binding of the first peptide is pY-dependent, but phosphorylation is not required for the second peptide. Our findings suggest a potential new function for the SH2 domain as a molecular clamp to promote dimerization of signaling proteins.

Publication types

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

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Crystallography, X-Ray
  • Ligands
  • Mice
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Peptide Fragments / chemistry*
  • Peptide Fragments / genetics
  • Peptide Fragments / metabolism*
  • Phosphorylation / genetics
  • Phosphotyrosine / chemistry
  • Phosphotyrosine / metabolism
  • Protein Binding / physiology
  • Protein Structure, Secondary / genetics
  • Protein Structure, Tertiary / genetics
  • Surface Plasmon Resonance
  • src Homology Domains / genetics
  • src Homology Domains / physiology*

Substances

  • Ligands
  • Peptide Fragments
  • Phosphotyrosine