Abstract：
Nerve agents pose significant threats to civilian and military populations. The reactivation of acetylcholinesterase (AChE) is critical in treating acute poisoning, but there is still lacking broad-spectrum reactivators, which presents a big challenge. Therefore, insights gained from the reactivation kinetic analysis and molecular docking are essential for understanding the behavior of reactivators towards intoxicated AChE. In this research, we present a systematic determination of the reactivation kinetics of three V agents-inhibited four human ChEs [(AChE and butyrylcholinesterase (BChE)) from either native or recombinant resources, namely, red blood cell (RBC) AChE, rhAChE, hBChE, rhBChE) reactivated by five standard oximes. We unveiled the effect of native and recombinant ChEs on the reactivation kinetics of V agents ex vitro, where the reactivation kinetics characteristic of Vs-inhibited BChE was reported for the first time. In terms of the inhibition type, all of the five oxime reactivators exhibited noncompetitive inhibition. The inhibition potency of these reactivators would not lead to the difference in the reactivation kinetics between native and recombinant ChE. Despite the significant differences between the native and recombinant ChEs observed in the inhibition, aging, and spontaneous reactivation kinetics, the reactivation kinetics of V agent-inhibited ChEs by oximes were less differentiated, which were supported by the ligand docking results. We also found differences in the reactivation efficiency between five reactivators and the phosphorylated enzyme, and molecular dynamic simulations can further explain from the perspectives of conformational stability, hydrogen bonding, binding free energies, and amino acid contributions. By Poisson-Boltzmann surface area (MM-PBSA) calculations, the total binding free energy trends aligned well with the experimental kr2 values.
摘要：
神经毒剂对平民和军事人口构成重大威胁。乙酰胆碱酯酶（AChE）的再活化在治疗急性中毒中至关重要，但是仍然缺乏广谱活化剂，这是一个巨大的挑战。因此,从再活化动力学分析和分子对接获得的见解对于了解再活化剂对中毒AChE的行为至关重要。在这项研究中,我们提出了从天然或重组资源中抑制三种V剂的四种人类ChEs[（AChE和丁酰胆碱酯酶（BChE））的再激活动力学的系统测定，即,红细胞（RBC）AChE，rhAChE,hBChE,rhBChE）被五种标准肟重新激活。我们揭示了天然和重组ChEs对体外V剂再激活动力学的影响，首次报道了Vs抑制的BChE的再激活动力学特征。就抑制类型而言，所有五种肟再激活剂均表现出非竞争性抑制作用。这些再激活剂的抑制效力不会导致天然和重组ChE之间的再激活动力学的差异。尽管在抑制中观察到的天然和重组ChEs之间存在显着差异，老化，和自发再激活动力学，肟对V剂抑制的ChEs的再活化动力学分化较小，这得到了配体对接结果的支持。我们还发现五种激活剂和磷酸化酶之间的再激活效率存在差异，分子动力学模拟可以从构象稳定性的角度进一步解释，氢键,结合自由能，和氨基酸的贡献。通过泊松-玻尔兹曼表面积(MM-PBSA)计算，总结合自由能趋势与实验kr2值吻合良好。
