基于模块化组装和酶诱导解组装的超灵敏荧光检测胰蛋白酶的研究
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- 英文论文题名：Ultrasensitive Fluorescent detection of Trypsin Based on Modular Assemblies and Enzyme-Triggered Disassembly
- 论文作者：刘祥 ; 贾兰 ; 陈松
- 英文论文作者：Xiang Liu ; Lan Jia ; Song Chen ; Department of Materials science ; Institute of materials science and engineering ; TaiYuan University of Technology
- 年：2016
- 作者机构：太原理工大学材料学院材料学系;
- 论文关键词：荧光检测 ; 胰蛋白酶 ; 模块化组装体
- 英文论文关键词：Fluorometric assay ; Trypsin ; Modular Assemblies
- 会议召开时间：2016-07-01
- 会议录名称：中国化学会第30届学术年会摘要集-第二十四分会：超分子组装与软物质材料
- 语种：中文
- 分类号：O657.3;O629.8
- 学会代码：ZGHY
- 会议名称：中国化学会第30届学术年会-第二十四分会 ; 超分子组装与软物质材料
- 会议地点：中国辽宁大连
- 主办单位：中国化学会
- 学会名称：中国化学会
- 页数：1
- 文件大小：188k
- 原文格式：D
- 会议级别：全国
摘要
本研究基于模块化策略建立了一种超灵敏的胰蛋白酶检测荧光体系。将作为胰蛋白酶底物的鱼精蛋白设计为受体单元,疏水染料作为荧光报告单元。在表面活性剂浓度低于其临界胶束浓度条件下,带正电的鱼精蛋白和阴离子表面活性剂十二烷基磺酸钠通过静电作用组装,组装体的疏水内腔可以将疏水染料包埋进去。因此,受体单元和荧光报告单元通过非共价作用相互靠近。随后在组装体中加入胰蛋白酶,使鱼精蛋白水解为短肽,诱导组装体解组装以及染料的释放,荧光强度降低,以此检测酶的活性。得到的胰蛋白酶检测限为0.044 ng mL~(-1)。检测体系应用了三种荧光染料:尼罗红、芘和香豆素6,组装体的模块化特性使体系可通过改变荧光染料的类型来调整发射波长和检测范围。
A simple and ultra-sensitive fluorometric assay method for trypsin detection is successfully established based on modular strategy. Natural protein protamine, the substrate of trypsin, was designed as receptor unit, and hydrophobic dye was used as reporter unit, which converted binding events into fluorescent signals. The positively charged protamine would form micellar-type assemblies with anionic surfactant sodium dodecyl sulfate, which concentration was lower than its critical micelle concentration. The combination should provide supramolecular assemblies with apolar interior that could sequester hydrophobic fluorescent dye molecules. Hence, the receptor and the reporter units were kept in close proximity by means of non-covalent interactions. Subsequent addition of trypsin into the assemblies system led to cleavage of protamine into short peptides, triggering the dissociation of the assemblies and the release of dye. A detection level down to 0.044 ng mL~(-1) for trypsin was obtained. The dynamic nature of the organized assemblies made it possible to tune the emission wavelength and detection range just by simply varying the type of fluorescent dye.
A simple and ultra-sensitive fluorometric assay method for trypsin detection is successfully established based on modular strategy. Natural protein protamine, the substrate of trypsin, was designed as receptor unit, and hydrophobic dye was used as reporter unit, which converted binding events into fluorescent signals. The positively charged protamine would form micellar-type assemblies with anionic surfactant sodium dodecyl sulfate, which concentration was lower than its critical micelle concentration. The combination should provide supramolecular assemblies with apolar interior that could sequester hydrophobic fluorescent dye molecules. Hence, the receptor and the reporter units were kept in close proximity by means of non-covalent interactions. Subsequent addition of trypsin into the assemblies system led to cleavage of protamine into short peptides, triggering the dissociation of the assemblies and the release of dye. A detection level down to 0.044 ng mL~(-1) for trypsin was obtained. The dynamic nature of the organized assemblies made it possible to tune the emission wavelength and detection range just by simply varying the type of fluorescent dye.
引文
[1]N.D.Rawlings and A.J.Barrett,Proteolytic Enzymes:Serine and Cysteine Peptidases,1994,244,19.
[2]G.W.Slysz,D.F.Lewis and D.C.Schriemer,J.Proteome Res.,2006,5,1959.
[3]R.P.Liang,X.C.Tian,P.Qiu and J.D.Qin,Analytical Chemistry,2014,86,9256.
[4]L.Chen,X.Fu and J.Li,Nanoscale,2013,5,5905.
[2]G.W.Slysz,D.F.Lewis and D.C.Schriemer,J.Proteome Res.,2006,5,1959.
[3]R.P.Liang,X.C.Tian,P.Qiu and J.D.Qin,Analytical Chemistry,2014,86,9256.
[4]L.Chen,X.Fu and J.Li,Nanoscale,2013,5,5905.