Molecular Toxinology Laboratory, Department of Molecular Biology and Biotechnology, Tezpur University, Napaam, Tezpur, Assam, India.
Molecular Toxinology Laboratory, Department of Molecular Biology and Biotechnology, Tezpur University, Napaam, Tezpur, Assam, India.
Toxicon. 2021 Nov;203:93-103. doi: 10.1016/j.toxicon.2021.09.026. Epub 2021 Oct 4.
Inefficacy and associated risks of current antivenom has raised the need for alternative approaches of snakebite management. Aptamers are one such alternative which is being pursued for therapeutic interventions as well as for design of diagnostic kits due to its high specificity. Present study focussed on designing and validating nucleic acid aptamers against snake venom PLA a hydrolytic enzyme present in all venomous snakes. The aptamers were designed by adding nucleic acid chain on the surface of Daboxin P, a major PLA enzyme of Daboia russelii venom. Binding characteristics of the aptamers were confirmed by docking to Daboxin P as well as acidic and basic PLAs from different snake species using in silico docking. The aptamers folded into different tertiary structures and bound to the active and Ca binding site of PLA enzymes. Molecular dynamics simulation analysis of Daboxin P-aptamer complexes showed that the complexes were stable in an aqueous environment. The electrophoretic mobility shift assay further confirmed the binding of the synthetic aptamers to Daboxin P and other snake venom PLA enzymes. The aptamers inhibited the sPLA activity with an IC value ranging between 0.52 μM and 0.77 μM as well as the anticoagulant activity of Daboxin P. The aptamers could also inhibit the PLA activity of Echis carinatus crude venom and anti-coagulant activity of Bungarus caeruleus crude venom, members of big four snakes. However, the aptamers didn't inhibit fibrinogenolytic or proteolytic activity of big four venom as well as the coagulation and hemolytic activities. Thus, aptamers can be rationally designed to inhibit the biochemical and biological activities of snake venom proteins.
当前抗蛇毒血清的无效性及其相关风险促使人们需要寻找替代的蛇伤管理方法。适体是一种替代方法，由于其高度特异性，它正被用于治疗干预和诊断试剂盒的设计。本研究专注于针对蛇毒 PLA 这种存在于所有毒蛇毒液中的水解酶设计和验证核酸适体。通过在 Daboia russelii venom 中主要 PLA 酶 Daboxin P 的表面添加核酸链来设计适体。通过对接 Dabooxin P 以及来自不同蛇种的酸性和碱性 PLA 酶，使用计算机对接来验证适体的结合特性。适体折叠成不同的三级结构，并与 PLA 酶的活性和 Ca 结合位点结合。Daboxin P-适体复合物的分子动力学模拟分析表明，该复合物在水相环境中稳定。电泳迁移率变动分析进一步证实了合成适体与 Daboxin P 和其他蛇毒 PLA 酶的结合。适体以 0.52 μM 和 0.77 μM 之间的 IC 值抑制 sPLA 活性以及 Daboxin P 的抗凝活性。适体还可以抑制 Echis carinatus 粗毒液的 PLA 活性和 Bungarus caeruleus 粗毒液的抗凝活性，这两种毒液均来自四大毒蛇。然而，适体并没有抑制四大毒蛇毒液的纤维蛋白溶酶或蛋白水解活性以及凝血和溶血活性。因此，可以合理设计适体来抑制蛇毒蛋白的生化和生物学活性。
