Epitopes prediction for microcystin-LR by molecular docking
Epitopes prediction for microcystin-LR by molecular docking
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通过分子对接预测微囊藻毒素-LR的表位
DOI：
10.1016/j.ecoenv.2021.112925
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发表时间：
2021-10-28
影响因子：
6.8
通讯作者：
Zhu, Hao
中科院分区：
文献类型：
--
作者：
Liu, Yuan;Liu, Shu;Zhu, Hao
微囊藻毒素-LR (MC-LR) 是全球最有害的蓝藻毒素之一。许多针对 MC-LR 的抗体已被产生并表征。然而，关于MC-LR表位的知识仍然有限。本研究的目的是分析MC-LR的表位并证明MC-LR与其抗体的结合模式。针对 MC-LR 产生的小鼠杂交瘤细胞系 (Mab5H1-3B3) 的可变基因已被克隆并组装成单链可变片段 (scFv)，然后在大肠杆菌 BL21 中可溶性表达。基于scFv，通过竞争性ELISA测定MC-LR的IC50和IC10分别为7.45 nM和0.30 nM。 scFv还与MC-RR和MC-YR显示出115%和112%的交叉反应性，与MC-LA显示出59%的交叉反应性。通过分子对接，论证了MC-LR与其scFv之间的结合模式。在MC-LR的Adda5残基的羰基与其scFv之间观察到氢键相互作用，并且Arg4残基的胍基和MC-LR的Adda5残基的苯基也参与相互作用。这些预测的表位得到了抗体交叉反应数据的支持。通过比较 MCLR scFv 的抗体信息学与其预测的互补位，VH-CDR1 对于 MC-LR 结合至关重要，并且可以通过 V kappa-CDR1 和 V kappa-CDR3 中的工程来调整其特异性。这些信息对于微囊藻毒素的半抗原设计或改善 MC-LR scFv 的体外特性很有用。
Microcystin-LR (MC-LR) is one of the most worldwide harmful cyanobacterial toxins. A lots of antibodies against MC-LR have been generated and characterized. However, the knowledge about the epitopes of MC-LR was still limited. The objective of this study was to analyze the epitopes of MC-LR and demonstrate the binding mode of MC-LR with its antibody. The variable genes of a mouse hybridoma cell line (Mab5H1-3B3) raised against MC-LR have been cloned and assembled in a single chain variable fragment (scFv), and then soluble expressed in E.coli BL21. Based on the scFv, the IC50 and IC10 for MC-LR were determined to be 7.45 nM and 0.30 nM by competitive ELISA. And the scFv also showed 115% and 112% cross-reactivities to MC-RR and MC-YR, and 59% to MC-LA. By molecular docking, the binding mode between MC-LR and its scFv was demonstrated. A hydrogen bond interaction was observed between the carbonyl group of Adda5 residue of MC-LR and its scFv, and the guanidyl group of Arg4 residue and phenyl group of Adda5 residue of MC-LR were also involved in the interaction. These predicted epitopes were supported by antibody cross-reactivity data. By comparing the antibody informatics of MCLR scFv with its predicted paratopes, VH-CDR1 was crucial for MC-LR binding, and its specificity could be tuned by engineering in V kappa-CDR1 and V kappa-CDR3. These information would be useful for the hapten design for microcystins or improving the properties of MC-LR scFv in vitro.