A substrate-driven allosteric switch that enhances PDI catalytic activity

Nat Commun. 2016 Aug 30:7:12579. doi: 10.1038/ncomms12579.

Abstract

Protein disulfide isomerase (PDI) is an oxidoreductase essential for folding proteins in the endoplasmic reticulum. The domain structure of PDI is a-b-b'-x-a', wherein the thioredoxin-like a and a' domains mediate disulfide bond shuffling and b and b' domains are substrate binding. The b' and a' domains are connected via the x-linker, a 19-amino-acid flexible peptide. Here we identify a class of compounds, termed bepristats, that target the substrate-binding pocket of b'. Bepristats reversibly block substrate binding and inhibit platelet aggregation and thrombus formation in vivo. Ligation of the substrate-binding pocket by bepristats paradoxically enhances catalytic activity of a and a' by displacing the x-linker, which acts as an allosteric switch to augment reductase activity in the catalytic domains. This substrate-driven allosteric switch is also activated by peptides and proteins and is present in other thiol isomerases. Our results demonstrate a mechanism whereby binding of a substrate to thiol isomerases enhances catalytic activity of remote domains.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allosteric Regulation / drug effects
  • Animals
  • Blood Platelets / drug effects
  • Blood Platelets / physiology
  • Catalytic Domain / drug effects
  • Disease Models, Animal
  • Drug Evaluation, Preclinical
  • Endoplasmic Reticulum / metabolism*
  • Enzyme Inhibitors / pharmacology*
  • Enzyme Inhibitors / therapeutic use
  • Healthy Volunteers
  • Humans
  • Hydrophobic and Hydrophilic Interactions / drug effects
  • Insulin / metabolism
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Platelet Aggregation / drug effects*
  • Protein Binding / drug effects
  • Protein Disulfide-Isomerases / antagonists & inhibitors
  • Protein Disulfide-Isomerases / chemistry
  • Protein Disulfide-Isomerases / metabolism*
  • Protein Folding*
  • Protein Structure, Tertiary / drug effects
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism
  • Thrombosis / blood
  • Thrombosis / drug therapy
  • Thrombosis / pathology

Substances

  • Enzyme Inhibitors
  • Insulin
  • Recombinant Proteins
  • Protein Disulfide-Isomerases