原子力显微镜拉伸技术揭示金属蛋白酶ADAMTS13的构象  

Atomic Force Microscopy Pulling Reveals the Conformation of Metalloproteinase ADAMTS13

作　　者：谷慧婷 杨俊贤 方金花 吴建华[1] 林蒋国 GU Huiting;YANG Junxian;FANG Jinhua;WU Jianhua;LIN Jiangguo(School of Bioscience and Bioengineering,South China University of Technology,Guangzhou 510006,China;Research Department of Medical Sciences,Guangdong Provincial People’s Hospital(Guangdong Academy of Medical Sciences),Southern Medical University,Guangzhou 510080,China)

机构地区：[1]华南理工大学生物科学与工程学院,广州510006 [2]南方医科大学附属广东省人民医院(广东省医学科学院)医学研究中心,广州510080

出　　处：《医用生物力学》2025年第2期441-448,共8页Journal of Medical Biomechanics

基　　金：国家自然科学基金项目(32071303,12302400);广东省基础与应用基础研究基金(2023A1515012606)。

摘　　要：目的探究不同pH条件下ADAMTS13的构象态。方法利用原子力显微镜对两种拉伸体系(Biotin-Streptavidin体系和6×His-Anti-His抗体体系)下的ADAMTS13分子进行拉伸,并分析断裂力和分子轮廓长度。结果pH 7.4条件下,利用Biotin-Streptavidin体系拉伸ADAMTS13时,分子轮廓长度平均值为(30.93±1.56)nm,其频数分布为双峰分布,峰值位置分别为(22.12±0.01)、(49.57±0.05)nm;利用6×His-Anti-His抗体体系拉伸ADAMTS13时,分子轮廓长度平均值为(32.77±0.72)nm,其频数分布为双峰分布,峰值位置分别为(25.73±0.16)、(43.84±0.63)nm。pH 6.0条件下,利用Biotin-Streptavidin体系拉伸ADAMTS13时,分子轮廓长度平均值为(47.07±1.6)nm,其频数分布为3峰分布,峰值位置分别是(22.00±1.25)、(55.09±2.62)、(76.69±3.06)nm。与pH 7.4相比,pH 6.0时ADAMTS13的构象更为伸展。结论ADAMTS13在pH 7.4时存在“闭合”构象态和中间构象态;而在pH 6.0时,ADAMTS13存在“闭合”构象态、中间构象态和“开放”构象态。研究结果有助于进一步理解ADAMTS13在正常生理和血栓性血小板减少症紫癜中的作用,为新型重组ADAMTS13药物的开发提供思路。Objective To investigate the conformational states of ADAMTS13 under different pH conditions.Methods Atomic force microscopy(AFM)was applied to pull ADAMTS13 molecules with two distinct pulling systems:the Biotin-Streptavidin system and the 6×His-Anti-His antibody system.The rupture forces and molecular contour lengths were analyzed.Results Under pH 7.4 condition,when ADAMTS13 was pulled by Biotin-Streptavidin system,the mean molecular contour length was(30.93±1.56)nm,exhibiting a bimodal frequency distribution with peak positions at(22.12±0.01)and(49.57±0.05)nm.When ADAMTS13 was pulled using 6×Hist-anti-His antibody system,the mean molecular contour length was(32.77±0.72)nm,also showing a bimodal distribution,with a peak position at(25.73±0.16)and(43.84±0.63)nm,respectively.Under pH 6.0 condition,when ADAMTS13 was pulled by Biotin-Streptavidin system,the mean molecular contour length increased to(47.07±1.6)nm,and the frequency distribution shifted to trimodal,with peak positions at(22±1.25),(55.09±2.62)and(76.69±3.06)nm.The conformation of ADAMTS13 was more extended at pH 6.0 compared with that at pH 7.4.Conclusions ADAMTS13 exists in‘closed’and intermediate conformational states at physiological pH 7.4.However,at pH 6.0,ADAMTS13 can adopt‘closed’,intermediate,and‘open’conformational states.This study contributes to a further understanding of the role of ADAMTS13 in normal physiology and thrombotic thrombocytopenic purpura,providing insights for the development of novel recombination ADAMTS13 drugs.

关 键 词：ADAMTS13 原子力显微镜 构象状态 断裂力 分子轮廓长度 

分 类 号：R318.01[医药卫生—生物医学工程]