α-芋螺毒素LvIA特定氨基酸的突变对其活性的影响  

Effect of the mutation of specific amino acid onα-Conotoxin LvIA potency

作　　者：李浩楠 毛楷林 熊洋 长孙东亭[1] 朱晓鹏[1] 罗素兰[1] LI Hao-nan;MAO Kai-lin;XIONG Yang;ZHANGSUN Dong-ting;ZHU Xiao-peng;LUO Su-lan(Key Laboratory of Tropical Biological Resources,Ministry of Education,School of Life and Pharmaceutical Sciences,Key Lab for Marine Drugs of Haikou,Hainan University,Haikou 570228,China)

机构地区：[1]海南大学热带生物资源教育部重点实验室,生命科学与药学院,海口市海洋药物重点实验室,海南海口570228

出　　处：《中国海洋药物》2020年第6期45-51,共7页Chinese Journal of Marine Drugs

基　　金：海南省自然科学基金项目(818QN229);国家自然科学基金项目(81872794,31760249)资助。

摘　　要：目的对特异区分α6/α3β2β3和α3β22种nAChRs亚型的α-CTx LvIA进行研究,确定影响其活性的关键氨基酸位点。方法通过序列比对,两步法氧化折叠、双电极电压钳和圆二色谱等技术确定Lv IA中影响其活性的关键氨基酸位点,并探索该位点对Lv IA影响的机制。结果确定了Lv IA中第5位组氨酸(His,H)的关键作用,当第5位的His分别被丙氨酸(Ala,A)、天冬氨酸(Asp,D)和色氨酸(Trp,W)分别取代后,都会导致Lv IA对所有受体亚型的活性丧失。圆二色检测结果表明该位点的变化对Lv IA的二级结构没有显著影响。结论第5位His是维持Lv IA活性的关键氨基酸残基,该氨基酸极性和侧链的变化均会导致Lv IA活性的丧失,但是对其二级结构影响较小。Lv IA中第5位氨基酸与nAChR的相互作用力是保证其活性的关键。该结论为基于Lv IA设计新型专一作用于乙酰胆碱受体的工具探针改造提供了重要信息。Objective To identify the key amino acid residue that affects the activity of theα-CTx LvIA,which is specific to distinguish the related two nAChRs subtypesα6/α3β2β3 andα3β2.Methods By sequence alignment,two-step oxidative folding,two-electrode voltage clamp,and circular dichroism,the key amino acid residue affecting the activity of LvIA was identified and the relationship of the key site with the potency of LvIA was explored.Results The vital role of the fifth histidine(His5,H5)in LvIA was identified.When the His5 was substituted with alanine(Ala,A),aspartic acid(Asp,D)and tryptophan(Trp,W),respectively,the activities of LvIA mutants for all nAChR subtypes were lost.The circular dichroism results showed that the change of this site had no significant effect on the secondary structure of LvIA.Conclusion The His5 was the key amino acid residue to maintain the activity of LvIA.Changes in the polarity and side chains of His5 could lead to the loss of potency but have little effect on its secondary structure.The interaction between the5 th amino acid and nAChR in LvIA was the key to its activity.This conclusion provided key information for further design novel probe tools based on LvIA which specifically targets different nAChR subtypes.

关 键 词：α-芋螺毒素LvIA 烟碱型乙酰胆碱受体 突变体 电生理 活性 

分 类 号：R931.77[医药卫生—生药学]