Evidence for the extended helical nature of polysaccharide epitopes. The 2.8 A resolution structure and thermodynamics of ligand binding of an antigen binding fragment specific for alpha-(2-->8)-polysialic acid

Biochemistry. 1995 May 23;34(20):6737-44. doi: 10.1021/bi00020a019.

Abstract

The antigen binding fragment from an IgG2a kappa murine monoclonal antibody with specificity for alpha-(2-->8)-linked sialic acid polymers has been prepared and crystallized in the absence of hapten. Crystals were grown by vapor diffusion equilibrium with 16-18% polyethylene glycol 4000 solutions. The structure was solved by molecular replacement methods and refined to a conventional R factor of 0.164 for data to 2.8 A. The binding site is observed to display a shape and distribution of charges that is complementary to that of the predicted conformation of the oligosaccharide epitope. A thermodynamic description of ligand binding has been compiled for oligosaccharides ranging in length from 9 to 41 residues, and the data for the largest ligand has been used in a novel way to estimate the size of the antigen binding site. A model of antigen binding is presented that satisfies this thermodynamic data, as well as a previously reported requirement of conformational specificity of the oligosaccharide. X-ray crystallographic and thermodynamic evidence are consistent with a binding site that accommodates at least eight sialic acid residues.

MeSH terms

  • Antibodies, Monoclonal / chemistry*
  • Antibodies, Monoclonal / metabolism
  • Binding Sites, Antibody
  • Crystallization
  • Crystallography, X-Ray
  • Epitopes / chemistry*
  • Epitopes / immunology
  • Haptens / chemistry
  • Immunoglobulin Fab Fragments / chemistry
  • Immunoglobulin Fab Fragments / metabolism
  • Immunoglobulin G / chemistry
  • Immunoglobulin G / metabolism
  • Macromolecular Substances
  • Models, Molecular
  • Peptide Fragments / chemistry*
  • Protein Structure, Secondary*
  • Sialic Acids / chemistry
  • Sialic Acids / immunology*
  • Thermodynamics

Substances

  • Antibodies, Monoclonal
  • Epitopes
  • Haptens
  • Immunoglobulin Fab Fragments
  • Immunoglobulin G
  • Macromolecular Substances
  • Peptide Fragments
  • Sialic Acids
  • polysialic acid