Inhibition of antithrombin by Plasmodium falciparum histidine-rich protein II

Abstract

Histidine-rich protein II (HRPII) is an abundant protein released into the bloodstream by Plasmodium falciparum, the parasite that causes the most severe form of human malaria. Here, we report that HRPII binds tightly and selectively to coagulation-active glycosaminoglycans (dermatan sulfate, heparan sulfate, and heparin) and inhibits antithrombin (AT). In purified systems, recombinant HRPII neutralized the heparin-catalyzed inhibition of factor Xa and thrombin by AT in a Zn 2؉ -dependent manner. The observed 50% inhibitory concentration (IC 50 ) for the HRPII neutralization of AT activity is approximately 30nM for factor Xa inhibition and 90nM for thrombin inhibition. Zn 2؉ was required for these reactions with a distribution coefficient (K d ) of approximately 7M. Substituting Zn 2؉ with Cu 2؉ , but not with Ca 2؉ , Mg 2؉ , or Fe 2؉ , maintained the HRPII effect. HRPII attenuated the prolongation in plasma clotting time induced by heparin, suggesting that HRPII inhibits AT activity by preventing its stimulation by heparin. In the microvasculature, where erythrocytes infected with P falciparum are sequestered, high levels of released HRPII may bind cellular glycosaminoglycans, prevent their interaction with AT, and thereby contribute to the procoagulant state associated with P falciparum infection. (Blood. 2011;117(23): 6347-6354) IntroductionMalaria is an ancient parasitic disease that continues to take an enormous toll on persons and communities, as well as on global economies. In humans, the disease is caused by 5 protozoan parasites of the genus Plasmodium, including P ovale, P malariae, P vivax, P knowlesi, and P falciparum. Current infection rates are estimated at 250 million annually, with approximately 1 million of those cases resulting in fatalities. 1,2 P falciparum infection is the most dreaded, accounting for almost all malaria deaths. Morbidity and mortality from P falciparum malaria result from serious complications that arise during the course of the infection, including severe anemia, neurologic impairment, acute renal failure, and coagulopathy. Coagulation disturbances may contribute to the inflammation and organ failure associated with severe disease. 3,4 Patients with P falciparum infections have prolonged prothrombin time and activated partial thromboplastin time (aPTT), 5,6 reduced levels of clotting factors, including antithrombin (AT), 7,8 and elevated levels of fibrin degradation products. 9,10 Although uncommon, frank disseminated intravascular coagulation is observed in severe cases. 5,9,11 Activation of the intrinsic coagulation cascade and induction of tissue factor expression have been proposed, [12][13][14] but the precise mechanisms leading to hemostatic alterations in P falciparum infection remain poorly understood. Whether secreted parasite products are involved has not been established. Products released by intraerythrocytic parasites include a pair of proteins with unusually high histidine content: histidine-rich protein II (HRPII) and HRPIII. 15 Of these 2 homologous proteins...

Defibrotide Interferes With Several Steps of the Coagulation-Inflammation Cycle and Exhibits Therapeutic Potential to Treat Severe Malaria

“…Consumption of antihemostatics, 1 decrease in nitric oxide (NO) bioavailability, 2 distinct polymorphisms, 10 contribution of parasite-derived proteins ( e.g. HRPII) 11 and lipids ( e.g. Pf -GPI), 12 and events yet to be identified, may predispose to decompensation in cerebral and placental malaria.…”

Transcriptional variation in the malaria parasitePlasmodium falciparum

“…Genome Research 929 www.genome.org tivity (hrp) (Ndonwi et al 2011); and proteins located at the parasite-host cell interface (etramp) ( Table 1; Spielmann et al 2003). Surprisingly, many tRNAs were also expressed at different levels between different parasite lines.…”

Emerging roles for hemostatic dysfunction in malaria pathogenesis

“…84 Although the specific purpose of HRPII is not well understood, recent studies indicate that it binds glycosaminoglycans (GAGs) with exceptionally high affinity in the presence of zinc or copper ions. 85 Consequently, HRPII-GAG binding significantly impedes GAG-mediated antithrombin inhibition of FXa and thrombin in plasma and reverses the anticoagulant activity of heparin. 85 The ability of HRPII to potently inhibit a crucial plasma anticoagulant process implies a potential mechanism by which P falciparum-encoded proteins can directly modulate coagulation.…”

“…85 Consequently, HRPII-GAG binding significantly impedes GAG-mediated antithrombin inhibition of FXa and thrombin in plasma and reverses the anticoagulant activity of heparin. 85 The ability of HRPII to potently inhibit a crucial plasma anticoagulant process implies a potential mechanism by which P falciparum-encoded proteins can directly modulate coagulation. Together, these findings suggest that P falciparum triggers coagulation activation through multiple different pathways that include induction of TF expression on microvascular ECs; the exposure of negatively charged PS on the surface of IEs, which enables assembly of the intrinsic tenase and prothrombinase complexes; and direct inhibition of key endogenous anticoagulants (Figure 2).…”