Differential heme-mediated modulation of Deinagkistrodon, Dispholidus, Protobothrops and Pseudonaja hemotoxic venom activity in human plasma

Abstract

Envenomation by vipers with hemotoxic enzymes continues to be a worldwide source of morbidity and mortality. The present work examined the effects of exposure of venom enzymes to carbon monoxide and O-phenylhydroxylamine, agents that modulate the biometal heme, by forming carboxyheme and metheme, respectively. Four venoms obtained from medically important, diverse snake venom found in Africa, Asia and Australia were analyzed. The species that had venom tested in human plasma with thrombelastography and heme modulating agents were Deinagkistrodon acutus, Protobothrops mucrosquamatus, Dispholidus typus and Pseudonaja textilis. These venoms varied four hundred-fold in potency (ng-µg/ml) to exert procoagulant effects on human plasma; further, there was species specific variability in venom inhibition after exposure to carboxyheme or metheme agents. Lastly, using a wide range of carbon monoxide concentrations, it was determined that the factor V component of P. textilis venom was likely inhibited before the factor X component. Further investigation using this thrombelastograph-based, venom "kinetomic" methodology involving heme modulation will demonstrate in time its laboratory and clinical utility.

Ruthenium, Not Carbon Monoxide, Inhibits the Procoagulant Activity of Atheris, Echis, and Pseudonaja Venoms

“…The key element of the paradigm that implicates CO as the mechanism behind the effects of CORMs is the determination that the inactivated releasing molecule (iRM), the portion of the CORM that remains after CO release, has no effect or a different effect on the system tested with the CORM compared to the anticipated CO effect. This laboratory has used this CORM-based paradigm for the past few years to demonstrate that CO inhibited the various procoagulant and anticoagulant activities of hemotoxic venoms and enzymes collected from dozens of snake and lizard species [1][2][3][4][5][6][7][8][9][10][11][12][13]. The particular CORM used was CORM-2 (tricarbonyldichlororuthenium (II) dimer) [1][2][3][4][5][6][7][8][9][10][11][12][13].…”

“…This laboratory has used this CORM-based paradigm for the past few years to demonstrate that CO inhibited the various procoagulant and anticoagulant activities of hemotoxic venoms and enzymes collected from dozens of snake and lizard species [1][2][3][4][5][6][7][8][9][10][11][12][13]. The particular CORM used was CORM-2 (tricarbonyldichlororuthenium (II) dimer) [1][2][3][4][5][6][7][8][9][10][11][12][13]. The presumed mechanism was that CO must be interacting with a cryptic heme group attached to the various venoms and enzymes or in some other way interacting with these diverse enzymes and venoms [1][2][3][4][5][6][7][8][9][10][11][12][13].…”

“…The particular CORM used was CORM-2 (tricarbonyldichlororuthenium (II) dimer) [1][2][3][4][5][6][7][8][9][10][11][12][13]. The presumed mechanism was that CO must be interacting with a cryptic heme group attached to the various venoms and enzymes or in some other way interacting with these diverse enzymes and venoms [1][2][3][4][5][6][7][8][9][10][11][12][13]. Thus, this potentially heme-based, CO-modulated paradigm of snake venom activity seemed plausible with the aforementioned paradigm of CORM-CO release-inert iRM interactions with target molecules.…”

“…While it is unreasonable to reassess all previous venoms inhibited by CORM-2 to determine if a Ru-based radical rather than CO was mediating the inhibition [1][2][3][4][5][6][7][8][9][10][11][12][13], assessing a few representative venoms would be of benefit. To this end, three procoagulant venoms derived from diverse species from Africa and Australia were selected that have already been characterized as inhibited by CORM-2 but not by its iRM by this laboratory [8,10]. The species chosen are displayed in Table 1, and the venom proteomes of these particular and snakes within the same genus are similar in terms of presence of snake venom serine proteases (SVSP), snake venom metalloproteinases (SVMP), and PLA 2 [17][18][19][20][21].…”

“…The species chosen are displayed in Table 1, and the venom proteomes of these particular and snakes within the same genus are similar in terms of presence of snake venom serine proteases (SVSP), snake venom metalloproteinases (SVMP), and PLA 2 [17][18][19][20][21]. Fortuitously, archived aliquots of these three venoms that were never thawed or used in the original studies [8,10] were maintained at −80 • C and were available for the present investigation to test the hypothesis that inhibition by ruthenium molecular species and not carbon monoxide may be the mechanism by which these procoagulant venoms were inhibited by CORM-2. Considering the aforementioned, the present investigation had the following goals.…”