Department of Anesthesiology, University of Arizona College of Medicine, Tucson, AZ 85719, USA.
Int J Mol Sci. 2019 Sep 30;20(19):4853. doi: 10.3390/ijms20194853.
Snake venom enzymes of the L-amino acid oxidase (LAAO) class are responsible for tissue hemorrhage, edema, and derangement of platelet function. However, what role, if any, these flavoenzymes play in altering plasmatic coagulation have not been well defined. Using coagulation kinetomic analyses (thrombelastograph-based), it was determined that the LAAO derived from venom displayed a procoagulant activity associated with weak clot strength (no factor XIII activation) similar to thrombin-like enzymes. The procoagulant activity was not modified in the presence of reduced glutathione, demonstrating that the procoagulant activity was likely due to deamination, and not hydrogen peroxide generation by the LAAO. Further, unlike the raw venom of the same species, the purified LAAO was not inhibited by carbon monoxide releasing molecule-2 (CORM-2). Lastly, exposure of the enzyme to phenylmethylsulfonyl fluoride (PMSF) resulted in the LAAO expressing anticoagulant activity, preventing contact activation generated thrombin from forming a clot. In sum, this investigation for the first time characterized the LAAO of a snake venom as both a fibrinogen polymerizing and an anticoagulant enzyme acting via oxidative deamination and not proteolysis as is the case with thrombin-like enzymes (e.g., serine proteases). Using this thrombelastographic approach, future investigation of purified enzymes can define their biochemical nature.
蛇毒中的 L-氨基酸氧化酶（LAAO）类酶可导致组织出血、水肿和血小板功能紊乱。然而，这些黄素酶在改变血浆凝血方面起到何种作用（如果有的话）尚未得到很好的定义。采用凝血动力学分析（基于血栓弹性描记图的分析），发现源自蛇毒的 LAAO 具有促凝活性，与类凝血酶酶相似，与弱凝块强度（无因子 XIII 激活）相关联。在还原型谷胱甘肽存在的情况下，该促凝活性没有改变，这表明该促凝活性可能归因于脱氨作用，而不是 LAAO 产生的过氧化氢。此外，与同一物种的原始毒液不同，纯化的 LAAO 不受一氧化碳释放分子-2（CORM-2）的抑制。最后，酶暴露于苯甲基磺酰氟（PMSF）中导致 LAAO 表达抗凝活性，防止接触激活产生的凝血酶形成凝块。总之，这项研究首次对蛇毒的 LAAO 进行了特征描述，它既是纤维蛋白原聚合酶，也是抗凝酶，其作用机制通过氧化脱氨，而不是类凝血酶酶（例如丝氨酸蛋白酶）那样的蛋白水解作用。通过这种血栓弹性描记图的方法，未来对纯化酶的研究可以确定其生化性质。
