A synthetic camel anti-lysozyme peptide antibody (peptibody) with flexible loop structure identified by high-resolution affinity mass spectrometry

Chemistry. 2006 Feb 20;12(7):1915-23. doi: 10.1002/chem.200500785.


We describe the synthesis and characterisation of the fully functional molecular recognition structure of a 26-amino acid residue peptide antibody, referred to as peptibody, designed from a monoclonal single-domain antibody fragment derived from a camel heavy-chain antibody. The CDR3 region (CDR = complementarity determining region) of the cAbLys3 camel antibody fragment, which binds to the active site of hen eggwhite lysozyme (HEL) and acts as a potent enzyme inhibitor by mimicking an oligosaccharide substrate, was prepared by solid-phase peptide synthesis. To obtain a closed loop-like structure resembling that in the crystal structure, N- and C-terminal cysteine residues were added to the linear peptide and oxidised to a cyclic disulfide-bridged peptide by using dimethylsulfoxide. A further, internal cysteine-12 residue was acetamidomethyl-protected to prevent possible oxidative byproducts. Affinity separation on a lysozyme microcolumn combined with MALDI-TOF mass spectrometry revealed that the peptide resumed high affinity to lysozyme only after deprotection of Cys-12, suggesting the importance of this paratope sequence for epitope recognition. The complex of lysozyme and active peptibody was characterised directly by conducting high-resolution ESI-FTICR mass spectrometry, which provided a molecular comparison of affinities for linear and cyclic peptibodies.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Antibodies, Blocking / chemistry*
  • Antibodies, Blocking / pharmacology*
  • Camelus / immunology*
  • Chromatography, Affinity
  • Chromatography, High Pressure Liquid
  • Cyclization
  • Cysteine / chemistry
  • Linear Models
  • Mass Spectrometry
  • Models, Molecular
  • Molecular Sequence Data
  • Muramidase / antagonists & inhibitors*
  • Peptide Hydrolases / chemistry
  • Spectrometry, Mass, Electrospray Ionization
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization


  • Antibodies, Blocking
  • Muramidase
  • Peptide Hydrolases
  • Cysteine