Diblock copolymers enhance folding of a mechanosensitive membrane protein during cell-free expression
Diblock copolymers enhance folding of a mechanosensitive membrane protein during cell-free expression
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DOI：
10.1073/pnas.1814775116
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发表时间：
2019-03-05
影响因子：
11.1
通讯作者：
Kamat, Neha P.
中科院分区：
文献类型：
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作者：
Jacobs, Miranda L.;Boyd, Margrethe A.;Kamat, Neha P.
膜蛋白在囊泡膜中的表达和整合是设计拟细胞生物传感器、生物反应器和人工细胞的关键步骤。虽然膜蛋白已被整合到各种非天然膜中，但这些囊泡膜的化学和物理性质对蛋白质行为的影响在很大程度上仍然未知。非天然两亲物，如二嵌段共聚物，提供了一个界面，可以合成控制，以更好地研究这种关系。在这里，我们专注于膜蛋白生命周期的第一步：表达和折叠。我们观察到的模型mechanosensitive通道蛋白，mechanosensitive通道的大电导率，在无细胞反应过程中，当含有二嵌段共聚物的囊泡存在的折叠和整体生产的改进。通过系统地调整膜组合物的囊泡通过掺入的聚（环氧乙烷）-b-聚（丁二烯）二嵌段共聚物，我们表明，膜蛋白折叠和生产可以得到改善，在传统的脂质囊泡中观察到的。然后，我们用另一种膜弹性分子C12 E8重现这种效果。我们的研究结果表明，全球膜的物理性质，特别是可用的膜表面积和膜面积膨胀模量，显着影响膜蛋白的折叠和产量。此外，我们的研究结果为探索非天然膜两亲物如何用于研究和提高生物膜蛋白的生产奠定了基础。
The expression and integration of membrane proteins into vesicle membranes is a critical step in the design of cell-mimetic biosensors, bioreactors, and artificial cells. While membrane proteins have been integrated into a variety of nonnatural membranes, the effects of the chemical and physical properties of these vesicle membranes on protein behavior remain largely unknown. Nonnatural amphiphiles, such as diblock copolymers, provide an interface that can be synthetically controlled to better investigate this relationship. Here, we focus on the initial step in a membrane protein's life cycle: expression and folding. We observe improvements in both the folding and overall production of a model mechanosensitive channel protein, the mechanosensitive channel of large conductance, during cell-free reactions when vesicles containing diblock copolymers are present. By systematically tuning the membrane composition of vesicles through incorporation of a poly(ethylene oxide)-b-poly (butadiene) diblock copolymer, we show that membrane protein folding and production can be improved over that observed in traditional lipid vesicles. We then reproduce this effect with an alternate membrane-elasticizing molecule, C12E8. Our results suggest that global membrane physical properties, specifically available membrane surface area and the membrane area expansion modulus, significantly influence the folding and yield of a membrane protein. Furthermore, our results set the stage for explorations into how nonnatural membrane amphiphiles can be used to both study and enhance the production of biological membrane proteins.