基于分子模拟技术筛选宽体金线蛭抗凝活性肽  

An Anticoagulant Peptide Screened from Whitmania pigra Whitman Based on Molecular Simulation

作　　者：华羽彤 李尹 董瑞娟[1] 郭秀欢 雷艳 辛泉诚 魏鹏[1] 袁瑞娟[1] HUA Yu-tong;LI Yina;DONG Rui-juan;GUO Xiu-huan;LEI Yan;XIN Quan-cheng;WEI Peng;YUAN Rui-juan(Beijing University of Chinese Medicine,Beijing 102488,China)

机构地区：[1]北京中医药大学,北京102488

出　　处：《中国现代中药》2023年第9期1940-1948,共9页Modern Chinese Medicine

基　　金：国家自然科学基金项目(82173955);北京中医药大学校级科研纵向发展基金项目(2021-ZXFZJJ-070);北京中医药大学“青年科学家培育计划”项目;国家中医药管理局高水平中医药重点学科建设项目。

摘　　要：目的:采用分子模拟技术从宽体金线蛭来源多肽库中筛选抗凝活性多肽,结合分子对接初筛及分子动力学模拟复筛,最终通过抗凝实验验证得到宽体金线蛭抗凝活性多肽。方法:利用虚拟酶切技术对宽体金线蛭蛋白库进行酶切,得到宽体金线蛭多肽候选库;选择凝血酶为凝血活性靶点,采用HPEPDOCK及Discovery Studio CDOCKER两种方法进行分子对接,并利用分子动力学模拟技术对分子对接初筛结果中对接结合能高的复合物进行动力学模拟复筛,通过MM-PBSA模块分析计算凝血酶-多肽复合物结合自由能;最后采用固相合成法化学合成复筛中稳定结合的宽体金线蛭多肽并进行体外抗凝活性测定。结果:经过虚拟酶切共得到3317条无毒多肽,通过分子对接进行初筛,结合分子动力学模拟复筛,得到1条结合凝血酶效果最好的目标抗凝肽,该多肽序列为QNTVGLDDFFSSYER,结合自由能为-427.506 kJ·mol^(-1)。凝血酶Arg233残基是该凝血酶-多肽复合物中介导结合相互作用的关键氨基酸。该宽体金线蛭抗凝肽在体外活性测定中确能延长凝血酶时间。结论:利用分子模拟技术有效筛选出1条宽体金线蛭抗凝活性肽,并经过抗凝实验验证,表明“模拟+实验”的活性肽筛选策略切实可行,该研究思路可为动物类中药物质基础研究提供参考。Objective:To screen out anticoagulant peptides from the peptide library of Whitmania pigra Whitman by molecular docking and molecular dynamics simulation,and verify the predicted anticoagulant peptides by anticoagulation experiments.Methods:The in silico digestion method was used for the cleavage of W.pigra proteins,and a candidate peptide library was built.HPEPDOCK and Discovery Studio CDOCKER were employed to perform molecular docking,and molecular dynamics simulation was adopted to screen the peptide candidates capable of binding to the thrombin.MM-PBSA was used to calculate thrombin-peptide binding energy.Finally,the peptide with a stable binding activity was synthesized and its anticoagulant activity was measured in vitro.Results:After digestion,3317 non-toxic peptides were obtained,from which a peptide with the sequence QNTVGLDDFFSSYER was screened out,with the binding energy of–427.506 kJ·mol^(-1).The thrombin residue Arg233 may be the key amino acid for the interaction between thrombin and the peptide.The in vitro activity determination showed that this peptide prolonged the thrombin time.Conclusion:In this study,molecular simulation was employed to screen the anticoagulant peptide from W.pigra.The anticoagulation experiment verified that the strategy of simulation+experiment for the screening of active peptides was feasible.This study provides a new research idea for the screening of active ingredients from Chinese animal medicines.

关 键 词：分子对接 分子动力学模拟 宽体金线蛭 抗凝活性肽 

分 类 号：R285[医药卫生—中药学]