Liu Huipu, Chen Yunlong, Wang Jiawei, Yang Yuanjiao, Ju Huangxian
State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University Nanjing 210023 China
Chem Sci. 2021 Sep 2;12(43):14389-14395. doi: 10.1039/d1sc03059k. eCollection 2021 Nov 10.
Protein-membrane interactions play important roles in signal transductions and functional regulation of membrane proteins. Here, we design a molecular dynamometer (MDM) for analyzing protein-membrane interaction on living cells. The MDM is constructed by assembling an artificial lipid bilayer and alkylated Cy3-DNA azide (azide-Cy3-C) on a silica bubble. After a functional aptamer is covalently anchored onto the corresponding target protein on a living cell through UV irradiation, azide-Cy3-C is conjugated with the aptamer through a click reaction to produce a "tug-of-war" between the MDM and the cell due to the buoyancy of the silica bubble. This induces the detachment of the protein from the cell membrane or the alkane terminal from the MDM enabling sub-piconewton embedding force measurement by changing the alkane chain length and simple fluorescence analysis. The successful analysis of embedding force variation of a protein on the cell membrane upon post-translational modifications demonstrates the practicability and expansibility of this method for mechanics-related research in biological systems.
蛋白质-膜相互作用在信号转导和膜蛋白的功能调节中起着重要作用。在此，我们设计了一种分子测力计（MDM），用于分析活细胞上的蛋白质-膜相互作用。MDM是通过在二氧化硅气泡上组装人工脂质双层和烷基化的Cy3-DNA叠氮化物（叠氮化物-Cy3-C）构建而成的。在通过紫外线照射将功能性适配体共价锚定到活细胞上的相应靶蛋白后，叠氮化物-Cy3-C通过点击反应与适配体缀合，由于二氧化硅气泡的浮力，在MDM和细胞之间产生“拔河”效应。这会导致蛋白质从细胞膜上脱离或烷烃末端从MDM上脱离，从而通过改变烷烃链长度和简单的荧光分析实现亚皮牛顿嵌入力的测量。对蛋白质在翻译后修饰后在细胞膜上嵌入力变化的成功分析证明了该方法在生物系统力学相关研究中的实用性和可扩展性。
