Directed discovery of bivalent peptide ligands to an SH3 domain

Protein Sci. 2004 Mar;13(3):626-32. doi: 10.1110/ps.03470504.


The Caenorhabditis elegans SEM-5 SH3 domains recognize proline-rich peptide segments with modest affinity. We developed a bivalent peptide ligand that contains a naturally occurring proline-rich binding sequence, tethered by a glycine linker to a disulfide-closed loop segment containing six variable residues. The glycine linker allows the loop segment to explore regions of greatest diversity in sequence and structure of the SH3 domain: the RT and n-Src loops. The bivalent ligand was optimized using phage display, leading to a peptide (PP-G(4)-L) with 1000-fold increased affinity for the SEM-5 C-terminal SH3 domain over that of a natural ligand. NMR analysis of the complex confirms that the peptide loop segment is targeted to the RT and n-Src loops and parts of the beta-sheet scaffold of this SH3 domain. This binding region is comparable to that targeted by a natural non-PXXP peptide to the p67(phox) SH3 domain, a region not known to be targeted in the Grb2 SH3 domain family. PP-G(4)-L may aid in the discovery of additional binding partners of Grb2 family SH3 domains.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adaptor Proteins, Signal Transducing*
  • Amino Acid Sequence
  • Animals
  • Binding Sites / genetics
  • Caenorhabditis elegans Proteins / chemistry
  • Caenorhabditis elegans Proteins / genetics
  • Caenorhabditis elegans Proteins / metabolism*
  • GRB2 Adaptor Protein
  • Ligands
  • Membrane Proteins / chemistry
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism
  • Models, Molecular
  • Molecular Sequence Data
  • Nuclear Magnetic Resonance, Biomolecular
  • Peptide Fragments / chemistry
  • Peptide Fragments / genetics
  • Peptide Fragments / metabolism
  • Peptide Library
  • Peptides / chemical synthesis
  • Peptides / chemistry
  • Peptides / metabolism*
  • Phosphoproteins / chemistry
  • Phosphoproteins / genetics
  • Phosphoproteins / metabolism
  • Protein Binding
  • Protein Structure, Secondary
  • Proteins / chemistry
  • Proteins / genetics
  • Proteins / metabolism
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Thermodynamics
  • src Homology Domains / genetics
  • src Homology Domains / physiology*


  • Adaptor Proteins, Signal Transducing
  • Caenorhabditis elegans Proteins
  • GRB2 Adaptor Protein
  • Ligands
  • Membrane Proteins
  • Peptide Fragments
  • Peptide Library
  • Peptides
  • Phosphoproteins
  • Proteins
  • Recombinant Proteins
  • Sem-5 protein, C elegans
  • neutrophil cytosol factor 67K