Department of Biotechnology and Biomedicine, Technical University of Denmark, Produktionstorvet, Building 423, 2800, Kgs. Lyngby, Denmark.
Novo Nordisk A/S, Brennum Park 24 K, 3400, Hillerød, Denmark.
Anal Bioanal Chem. 2020 Sep;412(24):6307-6318. doi: 10.1007/s00216-020-02533-8. Epub 2020 Mar 12.
Vesicles constructed of either synthetic polymers alone (polymersomes) or a combination of polymers and lipids (lipo-polymersomes) demonstrate excellent long-term stability and ability to integrate membrane proteins. Applications using lipo-polymersomes with integrated membrane proteins require suitable supports to maintain protein functionality. Using lipo-polymersomes loaded with the light-driven proton pump bacteriorhodopsin (BR), we demonstrate here how the photocurrent is influenced by a chosen support. In our study, we deposited BR-loaded lipo-polymersomes in a cross-linked polyelectrolyte multilayer assembly either directly physisorbed on gold electrode microchips or cross-linked on an intermediary polyethersulfone (PES) membrane covalently grafted using a hydrogel cushion. In both cases, electrochemical impedance spectroscopic characterization demonstrated successful polyelectrolyte assembly with BR-loaded lipo-polymersomes. Light-induced proton pumping by BR-loaded lipo-polymersomes in the different support constructs was characterized by amperometric recording of the generated photocurrent. Application of the hydrogel/PES membrane support together with the polyelectrolyte assembly decreased the transient current response upon light activation of BR, while enhancing the generated stationary current to over 700 nA/cm. On the other hand, the current response from BR-loaded lipo-polymersomes in a polyelectrolyte assembly without the hydrogel/PES membrane support was primarily a transient peak combined with a low-nanoampere-level stationary photocurrent. Hence, the obtained results demonstrated that by using a hydrogel/PES support it was feasible to monitor continuously light-induced proton flux in biomimetic applications of lipo-polymersomes. Graphical abstract.
由合成聚合物单独构建的囊泡（聚合物囊泡）或聚合物和脂质的组合（脂聚合物囊泡）表现出优异的长期稳定性和整合膜蛋白的能力。使用整合了膜蛋白的脂聚合物囊泡的应用需要合适的载体来维持蛋白质的功能。在这里，我们使用负载有光驱动质子泵菌视紫红质（BR）的脂聚合物囊泡演示了所选载体如何影响光电流。在我们的研究中，我们将负载有 BR 的脂聚合物囊泡沉积在交联聚电解质多层组件中，这些囊泡直接物理吸附在金电极微芯片上，或者交联在中间的聚醚砜（PES）膜上，该膜通过水凝胶垫共价接枝。在这两种情况下，电化学阻抗谱表征都证明了带有 BR 负载的脂聚合物囊泡的成功聚电解质组装。通过安培记录生成的光电流来表征不同载体结构中负载有 BR 的脂聚合物囊泡的光诱导质子泵作用。应用水凝胶/PES 膜载体与聚电解质组装一起，降低了 BR 光激活时瞬态电流响应，同时将生成的稳定电流增强至超过 700 nA/cm。另一方面，在没有水凝胶/PES 膜载体的聚电解质组装中，负载有 BR 的脂聚合物囊泡的电流响应主要是瞬态峰值，加上低纳安级别的稳定光电流。因此，所得结果表明，通过使用水凝胶/PES 支撑物，可以在脂聚合物囊泡的仿生应用中连续监测光诱导质子通量。
