title: A novel heparin-dependent inhibitor of activated protein C that potentiates consumptive coagulopathy in Russell's viper envenomation.
abstract: The activation of coagulation factors V and X by Russell's viper venom (RVV) has been implicated in the development of consumptive coagulopathies in severely envenomed patients. However, factor Va is prone to inactivation by activated protein C (APC), an important serine protease that negatively regulates blood coagulation. It is therefore hypothesized that APC may be down-regulated by some of the venom components. In this study, we managed to isolate a potent Kunitz-type APC inhibitor, named DrKIn-I. Using chromogenic substrate, DrKIn-I dose-dependently inhibited the activity of APC. Heparin potentiated the inhibition and reduced the IC(50) of DrKIn-I by 25-fold. DrKIn-I, together with heparin, also protected factor Va from APC-mediated inactivation. Using surface plasmon resonance, DrKIn-I exhibited fast binding kinetics with APC (association rate constant = 1.7 × 10(7) M(-1) s(-1)). Direct binding assays and kinetic studies revealed that this inhibition (K(i) = 53 pM) is due to the tight binding interactions of DrKIn-I with both heparin and APC. DrKIn-I also effectively reversed the anticoagulant activity of APC and completely restored the thrombin generation in APC-containing plasma. Furthermore, although the injection of either DrKIn-I or RVV-X (the venom factor X-activator) into ICR mice did not significantly deplete the plasma fibrinogen concentration, co-administration of DrKIn-I with RVV-X resulted in complete fibrinogen consumption and the deposition of fibrin thrombi in the glomerular capillaries. Our results provide new insights into the pathogenesis of RVV-induced coagulopathies and indicate that DrKIn-I is a novel APC inhibitor that is associated with potentially fatal thrombotic complications in Russell's viper envenomation.
authors.affiliation: Institute of Biological Chemistry, Academia Sinica, Taipei 11529, Taiwan.
authors.forename: An-Chun
authors.initials: AC
authors.lastname: Cheng
authors.forename: Hua-Lin
authors.initials: HL
authors.lastname: Wu
authors.forename: Guey-Yueh
authors.initials: GY
authors.lastname: Shi
authors.forename: Inn-Ho
authors.initials: IH
authors.lastname: Tsai
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001777:Blood Coagulation / Q000187:drug effects; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D004211:Disseminated Intravascular Coagulation / Q000139:chemically induced / Q000378:metabolism*; D004357:Drug Synergism; D006493:Heparin / Q000494:pharmacology*; D006801:Humans; D007700:Kinetics; D051379:Mice; D008813:Mice, Inbred ICR; D008969:Molecular Sequence Data; D011485:Protein Binding; D011486:Protein C / Q000037:antagonists & inhibitors* / Q000378:metabolism; D018046:Protein C Inhibitor / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D017840:Daboia / Q000235:genetics / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D014757:Viper Venoms / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology / Q000506:poisoning
chemical_list: D002352:Carrier Proteins; D018076:DNA, Complementary; C573935:DrKIn-I protein, Daboia russelii; D011486:Protein C; D018046:Protein C Inhibitor; D014757:Viper Venoms; D006493:Heparin