Characterization of micropatterned lipid membranes on a gold surface by surface plasmon resonance imaging and electrochemical signaling of a pore-forming protein.
Characterization of micropatterned lipid membranes on a gold surface by surface plasmon resonance imaging and electrochemical signaling of a pore-forming protein.
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通过表面等离子共振成像和成孔蛋白的电化学信号表征金表面上的微图案脂质膜。
DOI：
10.1021/la051937m
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发表时间：
2005
期刊：
影响因子：
--
通讯作者：
Q. Cheng
中科院分区：
文献类型：
--
作者：
Zhuangzhi Wang;T. Wilkop;Q. Cheng
我们报道了一种用于化脓性链球菌致孔毒素--链球菌溶血素O(SLO)电化学信号传递的微图案膜电极的制备和表征。利用烷基硫醇单分子层的微接触印刷技术制备了阵列模板，并将含胆固醇的DMPC囊泡融合到该模板上形成脂层结构。通过原位表面等离子体共振(SPR)成像和电化学表征了支撑膜的结构，包括图案转移和囊泡融合。通过SPR角位移测量对所得膜进行了定量分析，结果表明，亲水性口袋中的膜的平均厚度为8.2+/-0.4 nm。结合荧光显微镜研究，结果表明这可能是一个混合的脂类集合体，可能由双层、囊泡碎片和脂联组成。通过测定氧化还原探针二茂铁羧酸(FCA)的伏安响应来定量毒素对支撑膜的作用。电化学测试表明，模板上的囊泡融合阻止了FCA的进入，而注射SLO毒素则恢复了氧化还原反应。FCA的氧化峰电流随毒素浓度的增加而增大，在40HU/mL时达到平台值。该方法具有很高的灵敏度，0.1HU/mLSLO(1.25 PM)即可产生明确的响应。此外，它不需要在膜传感中使用高度绝缘层，这为开发膜靶向蛋白质和多肽的新型传感接口开辟了新的途径。
We report the fabrication and characterization of a micropatterned membrane electrode for electrochemical signaling of a bacterial pore-forming toxin, Streptolysin O (SLO) from S. pyogenes. Microcontact printing of an alkylthiol monolayer was used to fabricate an array template, onto which cholesterol-containing DMPC vesicles were fused to form lipid layer structures. The construction of the supported membranes, including pattern transfer and vesicle fusion, was characterized by in-situ surface plasmon resonance (SPR) imaging and electrochemistry. Quantitative analysis of the resulting membrane by using SPR angular shift measurements indicates that the membranes in the hydrophilic pockets have an average thickness of 8.2 +/- 0.4 nm. Together with fluorescence microscopy studies, the results suggest that this could be a mixed lipid assembly that may consist of a bilayer, vesicle fragments, and lipid junctions. The voltammetric response of the redox probe ferrocene carboxylic acid (FCA) was measured to quantify the toxin action on the supported membrane. The electrochemical measurements indicate that fusion of vesicles on the template blocked the access of FCA, whereas the injection of SLO toxin restored the redox response. The anodic peak current of FCA was found to increase with toxin concentration until a plateau was reached at 40 HU/mL. The method is highly sensitive such that 0.1 HU/mL of SLO (1.25 pM) can yield a well-defined response. In addition, it eliminates the need for a highly insulating layer in membrane sensing, which opens up new avenues in developing novel sensing interfaces for membrane-targeting proteins and peptides.