Structural basis for thrombin activation of a protease-activated receptor: inhibition of intramolecular liganding

Chem Biol. 2003 Nov;10(11):1033-41. doi: 10.1016/j.chembiol.2003.10.014.

Abstract

Protease-activated G protein-coupled receptors (PAR1-4) are tethered-ligand receptors that are activated by proteolytic cleavage of the extracellular domain (exodomain) of the receptor. PAR1, the prototypic member of the PAR family, is the high-affinity thrombin receptor of platelets and vascular endothelium and plays a critical role in blood coagulation, thrombosis, and inflammation. Here, we describe the solution structure of the thrombin-cleaved exodomain of PAR1. The side chains of a hydrophobic hirudin-like (Hir) sequence and adjacent anionic motif project into solution. Docking of the exodomain Hir sequence to exosite I of thrombin reveals that the tethered ligand in the cleaved exodomain bends away from thrombin, leaving its active site available to another large macromolecular substrate. The N-terminal ligand is longer than anticipated and forms an intramolecular complex with a region located in the C terminus of the exodomain. Mutational analysis confirmed that this C-terminal region is a ligand binding site for both intra- and intermolecular ligands. A lipidated-ligand binding site peptide was found to be an effective inhibitor of thrombin-induced platelet aggregation.

Publication types

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

MeSH terms

  • Amino Acid Sequence
  • Ligands
  • Models, Molecular
  • Molecular Sequence Data
  • Nuclear Magnetic Resonance, Biomolecular
  • Protein Conformation
  • Receptors, Proteinase-Activated / antagonists & inhibitors
  • Receptors, Proteinase-Activated / metabolism*
  • Sequence Homology, Amino Acid
  • Thrombin / chemistry
  • Thrombin / metabolism*

Substances

  • Ligands
  • Receptors, Proteinase-Activated
  • Thrombin