Hemorrhagic toxins from western diamondback rattlesnake (Crotalus atrox) venom: isolation and characterization of five toxins and the role of zinc in hemorrhagic toxin e

Abstract

Five previously unknown hemorrhagic proteins, designated hemorrhagic toxins a,b,c,d, and e, were isolated from the venom of the western diamondback rattlesnake (Crotalus atrox). Molecular weights of hemorrhagic toxins a-e were determined to be 68 000, 24 000, 24 000, 24 000, and 25 700, respectively, by sodium dodecyl sulfate-phosphate gel electrophoresis using various polyacrylamide gel concentrations. Amino acid composition showed a total of 636, 200, 213, 214, and 219 amino acids for hemorrhagic toxins a-e, respectively. All the hemorrhagic toxins were found to lose their hemorrhagic activities with the metal chelators ethylenediaminetetraacetic acid and 1, 10-phenanthroline. All the hemorrhagic toxins were found to contain approximately 1 mol of zinc/mol of toxin, and they were all demonstrated to be proteolytic when dimethylcasein and dimethylhemoglobin were used as substrates. When zinc was removed from hemorrhagic toxin e with 1,10-phenanthroline, both both the proteolytic and hemorrhagic activities were equally inhibited. When the apohemorrhagic toxin e thus produced was incubated with zinc, the hemorrhagic and proteolytic activities were regenerated to the same extent. CD, UV, and Raman spectroscopy were used to study the structure of native hemorrhagin toxin e as well as the structural changes caused by zinc removal. From CD spectroscopy the native toxin was estimated to consist of 23% alpha helix, 6% beta structure, and 71% random-coil conformation. When over 90% of the zinc was removed, the alpha-helix content dropped from 23 to 7%.

Anti-hemolytic and Anti-cytotoxic Effect of Two Artemisia Species (A. campestris and A. herba-alba) Essential Oil against Snake Venom

“…Studies focused in some species to investigate and isolate the bioactive compounds from different part of the plant. The extracts from different parts of the plants belonging to the Asteraceae family showed an antihemorrhagic, antiproteolytic, anti-phospholipase (Abid et al, 2007;Mariane et al, 2011), antilethality (Shirwaikar et al, 2004;Pithayanukul et al, 2009), antiedema, (Ticli et al, 2005;Nishijima et al, 2009), antinecrosis (Bjarnason et al, 1978;Houghton et al, 1992;Chareanchai et al, 2009;Soares et al, 2005), anti-myotoxicity (Theakston and Reid, 1992;Bjarnason and Fox, 1994;Soares et al, 2004) and anticoagulant effect (Bjarnason and Tu, 1978;Bjarnason and Fox, 1994). Our results have some similarities with those research investigations.…”

Differential Evolution and Neofunctionalization of Snake Venom Metalloprotease Domains

“…Although it is unclear which domain of multidomain SVMP is responsible for the hemorrhagic effect of this toxin, chelation of the Zn2ϩ ion in the protease domain inhibits this activity. It is likely, therefore, that the protease domain of multidomain SVMP is responsible for the hemorrhaging often observed in envenomations by snakes with venoms rich in this toxin type (54,55). Moreover, it is hypothesized that the presence of the cysteine-rich and disintegrin-like domains results in the increased potency of some forms of SVMP (P-III) in comparison with those that lack them (P-I and P-II).…”

Structural interaction of natural and synthetic inhibitors with the venom metalloproteinase, atrolysin C (form d).

“…Collagens (e.g., types I, 1, III, and X) are characterized as tightly wound triple helices; each chain of the triple helix in type IV collagen has [21][22][23][24] interruptions of the Gly-Xaa-Yaa repeat (41), which are generally more sensitive to proteolysis; Yaa is usually Pro or hydroxyproline. Collagenases usually cleave a specific GlyXaa bond, with Xaa a hydrophobic residue at S1' (35); this is confirmed (32) for stromelysin (MMP-3), where model compound assays probed the pronounced hydrophobic nature of the extended binding site region (S3-S2').…”