Champagne Patrick S, Seneci Lorenzo, Fry Bryan G
Adaptive Biotoxicology Lab, School of the Environment, University of Queensland, St. Lucia, QLD 4072, Australia.
Toxins (Basel). 2025 Aug 8;17(8):398. doi: 10.3390/toxins17080398.
Venoms of the Palearctic vipers in the genus cause severe procoagulant clinical effects, yet the precise molecular targets remain incompletely defined. To fill this toxicological knowledge gap, we tested five venoms-, , (Turkmenistan and Uzbekistan localities), and -using plasma clotting assays, Factors VII, X, XI, and XII and prothrombin zymogen activation assays, and SDS-PAGE to visualise Factor V (FV) cleavage. All venoms induced extremely rapid clot formation (10.5-12.5 s) compared with the negative control (spontaneous clotting) of 334.6 ± 3.6 s) and the positive control (kaolin trigger) of 55.8 ± 1.9 s. Activation of FVII or FXI was negligible, whereas consistent FX activation and species-variable FXII activation, both moderate, were observed. Prothrombin remained inert in the absence of cofactors, but the presence of FV or FVa elicited potent thrombin generation. SDS-PAGE confirmed proteolytic conversion of the 330 kDa FV zymogen into the ~105 kDa heavy and ~80 kDa light chains of FVa by the venoms of all species. This data demonstrates that venoms rely on a dual enzyme strategy: (i) activation of FV to FVa by serine proteases and (ii) FVa-dependent prothrombin activation by metalloproteases. These results reveal that prothrombin activation is the dominant procoagulant pathway and overshadows the historically emphasised FX activation. This mechanism mirrors, yet is evolutionarily independent from, the FXa:FVa prothrombinase formation seen in Australian elapid venoms, highlighting convergent evolution of cofactor-hijacking strategies among snakes. The discovery of potent FVa-mediated prothrombin activation in challenges existing paradigms of viperid venom action, prompts re-evaluation of related genera (e.g., ), and underpins the design of targeted antivenom and therapeutic interventions.
该属古北蝰蛇的毒液会引发严重的促凝血临床效应，但确切的分子靶点仍未完全明确。为填补这一毒理学知识空白，我们使用血浆凝血试验、凝血因子VII、X、XI和XII以及凝血酶原酶原激活试验，并通过SDS-PAGE来观察凝血因子V（FV）的裂解情况，对五种毒液—— 、 、 （土库曼斯坦和乌兹别克斯坦产地）以及 进行了测试。与阴性对照（自发凝血，334.6 ± 3.6秒）和阳性对照（高岭土触发，55.8 ± 1.9秒）相比，所有毒液均诱导出极快速的凝血形成（10.5 - 12.5秒）。FVII或FXI的激活可忽略不计，而观察到一致的FX激活以及适度的、物种可变的FXII激活。在没有辅因子的情况下，凝血酶原保持惰性，但FV或FVa的存在会引发有效的凝血酶生成。SDS-PAGE证实，所有物种的毒液均将330 kDa的FV酶原蛋白水解转化为105 kDa的重链和80 kDa的FVa轻链。这些数据表明， 毒液依赖于双重酶策略：（i）丝氨酸蛋白酶将FV激活为FVa，以及（ii）金属蛋白酶依赖FVa激活凝血酶原。这些结果表明，凝血酶原激活是主要的促凝血途径，使历史上强调的FX激活黯然失色。这种机制与澳大利亚眼镜蛇科毒液中所见的FXa:FVa凝血酶原酶形成相似，但在进化上独立，突出了蛇类之间辅因子劫持策略的趋同进化。在 中发现强效的FVa介导的凝血酶原激活，挑战了蝰蛇毒液作用的现有范式，促使对相关属（如 ）进行重新评估，并为靶向抗蛇毒血清和治疗干预措施的设计提供了依据。
