A common site within factor H SCR 7 responsible for binding heparin, C-reactive protein and streptococcal M protein

Eur J Immunol. 2003 Apr;33(4):962-9. doi: 10.1002/eji.200323541.

Abstract

The complement inhibitor factor H (fH) interacts via its seventh short consensus repeat (SCR) domain with multiple ligands including heparin, streptococcal M protein and C-reactive protein (CRP). The aim of this study was to localize the residues in SCR 7 required for these interactions. We initially built a homology model of fH SCR 6-7 using the averaged NMR structures of fH SCR 15-16 and vaccinia control protein SCR 3-4 as templates. Electrostatic potentials of the model's surface demonstrated a co-localization of three clusters of positively charged residues on SCR 7, labeled site A (R369 and K370), site B (R386 and K387) and site C (K392). These residues, localized to the linker region preceding SCR 7 and to the end of a "hypervariable loop" in SCR 7, were systematically replaced with uncharged alanine residues in an fH construct containing SCR 1-7. The resulting proteins were expressed in the methylotrophic yeast, Pichia pastoris. By ELISA analysis we demonstrated: first, that substituting site A inhibited heparin and CRP binding; secondly, that substituting site B inhibited binding to heparin, CRP and M protein; and thirdly, that substituting site C clearly inhibited only heparin binding.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Substitution
  • Antigens, Bacterial*
  • Bacterial Outer Membrane Proteins / metabolism*
  • Binding Sites
  • C-Reactive Protein / metabolism*
  • Carrier Proteins / metabolism*
  • Complement Factor H / chemistry*
  • Complement Factor H / genetics
  • Complement Factor H / metabolism*
  • Heparin / metabolism*
  • Humans
  • Models, Molecular
  • Mutagenesis, Site-Directed
  • Protein Structure, Tertiary
  • Repetitive Sequences, Amino Acid
  • Structural Homology, Protein

Substances

  • Antigens, Bacterial
  • Bacterial Outer Membrane Proteins
  • Carrier Proteins
  • complement factor H, human
  • streptococcal M protein
  • Complement Factor H
  • Heparin
  • C-Reactive Protein