Selection of active ScFv to G-protein-coupled receptor CCR5 using surface antigen-mimicking peptides

Biochemistry. 2004 Oct 5;43(39):12575-84. doi: 10.1021/bi0492152.


This study describes the use of cyclic peptides for use in the selection of single-chain (ScFv) antibodies specific for the HIV-1 coreceptor CCR5, a representative G-protein-coupled receptor (GPCR). A tandem ligation strategy was developed for preparing biotinylated cyclic peptides, first through an orthogonal end-to-end ligation and then a chemoselective ligation with functionalized biotin. Cyclic peptides mimicking the extracellular loops of CCR5 and their unconstrained counterparts were then used for solution-phase selection of ScFv antibodies from a phage display antibody library. Antibodies reactive with CCR5 on cells were detected using a homogeneous high throughput assay. Of 19 isolated ScFv antibodies that bound to CCR5+ cells, three inhibited CCR5-mediated but not CXCR4-mediated HIV infection. Only ScFvs selected by binding to cyclic constrained peptides exhibited inhibitory activity. Our results demonstrate that surface-antigen mimetics of a GPCR are effective tools for selecting active, site-specific ScFv antibodies that hold promise as immunological reagents and therapeutics.

Publication types

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

MeSH terms

  • Amino Acid Sequence
  • Anti-HIV Agents / chemistry
  • Anti-HIV Agents / metabolism
  • Antigens, Surface / chemistry
  • Antigens, Surface / immunology
  • Antigens, Surface / metabolism*
  • Binding Sites, Antibody*
  • Cell Line, Tumor
  • Cross Reactions
  • Cysteine / chemistry
  • Enzyme-Linked Immunosorbent Assay
  • Esters
  • Growth Inhibitors / chemistry
  • Growth Inhibitors / metabolism
  • HIV-1 / growth & development
  • HIV-1 / immunology
  • HeLa Cells
  • Humans
  • Immunoglobulin Variable Region / chemistry
  • Immunoglobulin Variable Region / metabolism*
  • Ligands
  • Molecular Mimicry* / immunology
  • Molecular Sequence Data
  • Peptide Library
  • Peptides, Cyclic / chemical synthesis
  • Peptides, Cyclic / metabolism*
  • Receptors, CCR5 / chemistry
  • Receptors, CCR5 / immunology
  • Receptors, CCR5 / metabolism*
  • Sulfhydryl Compounds / chemistry


  • Anti-HIV Agents
  • Antigens, Surface
  • Esters
  • Growth Inhibitors
  • Immunoglobulin Variable Region
  • Ligands
  • Peptide Library
  • Peptides, Cyclic
  • Receptors, CCR5
  • Sulfhydryl Compounds
  • Cysteine