Mechanisms Responsible for the Anticoagulant Properties of Neurotoxic Dendroaspis Venoms: A Viscoelastic Analysis.

Vance G Nielsen,Nathaniel Frank,Michael T Wagner

doi:10.3390/ijms21062082

Vance G Nielsen, Nathaniel Frank + Show 1 more

Open Access

https://doi.org/10.3390/ijms21062082

Copy DOI

Abstract

Using thrombelastography to gain mechanistic insights, recent investigations have identified enzymes and compounds in Naja and Crotalus species’ neurotoxic venoms that are anticoagulant in nature. The neurotoxic venoms of the four extant species of Dendroaspis (the Black and green mambas) were noted to be anticoagulant in nature in human blood, but the mechanisms underlying these observations have never been explored. The venom proteomes of these venoms are unique, primarily composed of three finger toxins (3-FTx), Kunitz-type serine protease inhibitors (Kunitz-type SPI) and <7% metalloproteinases. The anticoagulant potency of the four mamba venoms available were determined in human plasma via thrombelastography; vulnerability to inhibition of anticoagulant activity to ethylenediaminetetraacetic acid (EDTA) was assessed, and inhibition of anticoagulant activity after exposure to a ruthenium (Ru)-based carbon monoxide releasing molecule (CORM-2) was quantified. Black mamba venom was the least potent by more than two orders of magnitude compared to the green mamba venoms tested; further, Black Mamba venom anticoagulant activity was not inhibited by either EDTA or CORM-2. In contrast, the anticoagulant activities of the green mamba venoms were all inhibited by EDTA to a greater or lesser extent, and all had anticoagulation inhibited with CORM-2. Critically, CORM-2-mediated inhibition was independent of carbon monoxide release, but was dependent on a putative Ru-based species formed from CORM-2. In conclusion, there was great species-specific variation in potency and mechanism(s) responsible for the anticoagulant activity of Dendroaspis venom, with perhaps all three protein classes—3-FTx, Kunitz-type SPI and metalloproteinases—playing a role in the venoms characterized.

Highlights

While the coagulopathic effects of hemotoxic venoms derived from venomous snakes have been of great scientific interest for several decades [1], it is only recently that investigation has intensified on the effects of neurotoxic venoms on coagulation [2,3,4,5,6,7]
As for the detection of a contribution of metalloproteinases to anticoagulant activity, venom derived from the Black Mamba was not affected by ethylenediaminetetraacetic acid (EDTA), suggesting that other venom constituents beyond SVMPs were responsible
The venoms of D. angusticeps and D. jamesoni had their anticoagulant activity extinguished by EDTA exposure, indicating that their anticoagulant activities were mediated by metalloproteinases exclusively

Summary

Introduction

While the coagulopathic effects of hemotoxic venoms derived from venomous snakes have been of great scientific interest for several decades [1], it is only recently that investigation has intensified on the effects of neurotoxic venoms on coagulation [2,3,4,5,6,7]. The venoms containing neurotoxic phospholipase A2 (PLA2) that were investigated were found to have their anticoagulant effects inhibited by either specific phospholipase A2 inhibitors or tricarbonyldichlororuthenium (II) dimer (CORM-2) [3,4,5,6,7] These preliminary investigations exploited the ability to assess changes in the hemostatic effects of enzyme-based neurotoxins in response to inhibitor exposure. As D. polylepis venom has a greater amount of Kunitz-type SPI than 3-FTx compared to the other three species, and given that isolated 3-FTx have acted as plasmatic anticoagulants [16,17], perhaps the predominance of 3-FTx in the other mamba species’ venom that was tested could explain the differences in anticoagulant potency [11,12]. Considered in composite, a few key proteins from these two non-enzymatic protein classes in combination with metalloproteinases could account for the venom anticoagulant activity and differences in anticoagulant potency observed in Dendroaspis species

Methods

Results

Conclusion