Inci Fatih, Celik Umit, Turken Basak, Özer Hakan Özgür, Kok Fatma Nese
Istanbul Technical University, Molecular Biology, Genetics and Biotechnology Program, 34469 Istanbul, Turkey.
Istanbul Technical University, Metallurgical and Materials Engineering Department, 34469 Istanbul, Turkey.
Biochem Biophys Rep. 2015 Jun 4;2:115-122. doi: 10.1016/j.bbrep.2015.05.012. eCollection 2015 Jul.
To investigate drug-membrane protein interactions, an artificial tethered lipid bilayer system was constructed for the functional integration of membrane proteins with large extra-membrane domains such as multi-drug resistance protein 1 (MDR1). In this study, a modified lipid (, 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino (polyethylene glycol)-2000] (DSPE-PEG)) was utilized as a spacer molecule to elevate lipid membrane from the sensor surface and generate a reservoir underneath. Concentration of DSPE-PEG molecule significantly affected the liposome binding/spreading and lipid bilayer formation, and 0.03 mg/mL of DSPE-PEG provided optimum conditions for membrane protein integration. Further, the incorporation of MDR1 increased the local rigidity on the platform. Antibody binding studies showed the functional integration of MDR1 protein into lipid bilayer platform. The platform allowed to follow MDR!-statin-based drug interactions . Each binding event and lipid bilayer formation was monitored in real-time using Surface Plasmon Resonance and Quartz Crystal Microbalance-Dissipation systems, and Atomic Force Microscopy was used for visualization experiments.
为了研究药物与膜蛋白的相互作用，构建了一种人工拴系脂质双层系统，用于膜蛋白与具有大的膜外结构域的蛋白（如多药耐药蛋白1，MDR1）的功能整合。在本研究中，一种修饰的脂质（1,2-二硬脂酰-sn-甘油-3-磷酸乙醇胺-N-[氨基（聚乙二醇）-2000]，DSPE-PEG）被用作间隔分子，以将脂质膜从传感器表面提升并在其下方形成一个储存库。DSPE-PEG分子的浓度显著影响脂质体的结合/铺展以及脂质双层的形成，0.03 mg/mL的DSPE-PEG为膜蛋白整合提供了最佳条件。此外，MDR1的掺入增加了平台上的局部刚性。抗体结合研究表明MDR1蛋白在脂质双层平台上实现了功能整合。该平台可用于追踪基于MDR1-他汀类药物的相互作用。使用表面等离子体共振和石英晶体微天平-耗散系统实时监测每次结合事件和脂质双层的形成，并使用原子力显微镜进行可视化实验。
