Structural basis of citrate-dependent and heparan sulfate-mediated cell surface retention of cobra cardiotoxin A3

J Biol Chem. 2005 Mar 11;280(10):9567-77. doi: 10.1074/jbc.M412398200. Epub 2004 Dec 6.

Abstract

Anionic citrate is a major component of venom, but the role of venom citrate in toxicity other than its inhibitory effect on the cation-dependent action of venom toxins is poorly understood. By immobilizing Chinese hamster ovary cells in microcapillary tubes and heparin on sensor chips, we demonstrated that heparan sulfate-mediated cell retention of the major cardiotoxin (CTX) from the Taiwan cobra, CTX A3, near membrane surfaces is citrate-dependent. X-ray determination of a CTX A3-heparin hexasaccharide complex structure at 2.4 A resolution revealed a molecular mechanism for toxin retention in which heparin-induced conformational changes of CTX A3 lead to citrate-mediated dimerization. A citrate ion bound to Lys-23 and Lys-31 near the tip of loop II stabilizes hydrophobic contact of the CTX A3 homodimer at the functionally important loop I and II regions. Additionally, the heparin hexasaccharide interacts with five CTX A3 molecules in the crystal structure, providing another mechanism whereby the toxin establishes a complex network of interactions that result in a strong interaction with cell surfaces presenting heparan sulfate. Our results suggest a novel role for venom citrate in biological activity and reveal a structural model that explains cell retention of cobra CTX A3 through heparan sulfate-CTX interactions.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cell Membrane / metabolism*
  • Citrates / pharmacology*
  • Cobra Cardiotoxin Proteins / chemistry*
  • Cobra Cardiotoxin Proteins / metabolism*
  • Elapid Venoms / chemistry
  • Elapidae
  • Heparitin Sulfate / pharmacology*
  • Models, Molecular
  • Molecular Sequence Data
  • Oligosaccharides / chemistry
  • Protein Conformation
  • Protein Structure, Secondary
  • Sequence Alignment
  • Sequence Homology, Amino Acid
  • Taiwan

Substances

  • Citrates
  • Cobra Cardiotoxin Proteins
  • Elapid Venoms
  • Oligosaccharides
  • Heparitin Sulfate

Associated data

  • PDB/1XT3