Hybrid vesicles composed of lipids and block copolymers hold promise for increasing liposome stability and providing a stable environment for membrane proteins. Recently we reported the successful functional reconstitution of the integral membrane protein cytochrome bo3 (ubiquinol oxidase) into hybrid vesicles composed of a blend of phospholipids and a block copolymer (PBd-PEO). We demonstrated that these novel membrane environments stabilise the enzymes' activity, prolonging their functional lifetime [Chem. Commun. 52 (2016) 11020-11023]. This approach holds great promise for applications of membrane proteins where enhanced durability, stability and shelf-life will be essential to creating a viable technology. Here we present a detailed account of our methods for membrane protein reconstitution into hybrid vesicles and discuss tips and challenges when using block copolymers compared to pure phospholipid systems that are more common materials for this purpose. We also extend the characterisation of these hybrid vesicles beyond what we have previously reported and show: (i) hybrid membranes are less permeable to protons than phospholipid bilayers; (ii) extended enzyme activity data is presented over a period of 500 days, which fully reveals the truly remarkable enhancement in functional lifetime that hybrid vesicles facilitate.

中文翻译：

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混合脂质聚合物囊泡中整合膜蛋白的重组方法，以增强功能耐久性

由脂质和嵌段共聚物组成的混合囊泡有望提高脂质体的稳定性并为膜蛋白提供稳定的环境。最近，我们报道了整合膜蛋白细胞色素 bo3（泛醇氧化酶）成功地功能重组为由磷脂和嵌段共聚物 (PBd-PEO) 的混合物组成的混合囊泡。我们证明了这些新的膜环境稳定了酶的活性，延长了它们的功能寿命 [Chem. 交流。52 (2016) 11020-11023]。这种方法对膜蛋白的应用具有很大的前景，在这些应用中，增强的耐久性、稳定性和保质期对于创造可行的技术至关重要。在这里，我们详细介绍了我们将膜蛋白重组为混合囊泡的方法，并讨论了使用嵌段共聚物与纯磷脂系统相比的技巧和挑战，纯磷脂系统是用于此目的的更常见材料。我们还将这些杂化囊泡的表征扩展到我们之前报道的范围之外，并表明：（i）杂化膜对质子的渗透性低于磷脂双层；(ii) 在 500 天的时间内提供了扩展的酶活性数据，这充分揭示了混合囊泡促进的功能寿命的真正显着提高。(i) 杂化膜对质子的渗透性低于磷脂双层；(ii) 在 500 天的时间内提供了扩展的酶活性数据，这充分揭示了混合囊泡促进的功能寿命的真正显着提高。(i) 杂化膜对质子的渗透性低于磷脂双层；(ii) 在 500 天的时间内提供了扩展的酶活性数据，这充分揭示了混合囊泡促进的功能寿命的真正显着提高。