DNA nanotechnology enabled high-precision membrane engineering

项目摘要 脂质双层膜在细胞中形成多种多样的动态结构，以提供重要功能，包括营养 吸收，废物管理，信号转导等。了解分子机制， 细胞产生并改变其膜结构的细胞是细胞生物学的中心任务。公 确定了膜的物理和化学性质调节它们与蛋白质的相互作用， 进而形成膜景观。然而，在如何实现这一目标方面， 蛋白质作用于不同的膜曲率和张力，特别是当膜结构和/或 膜-蛋白质相互作用是短暂的。使用重构膜的无细胞系统提供了 研究这种复杂的分子相互作用的有力方法。然而，仍然迫切需要一个 精确设计的平台，（1）精致地控制几何，生物化学和机械 和（2）容易允许膜的生物化学和结构表征， 相关蛋白质。利用DNA纳米技术，一种自下而上的方法， 具有可编程形状、刚度、化学修饰和运动的分子组装，我们建议 建立纳米工程膜，以弥合膜操作的技术差距，并解开 蛋白质介导的膜动力学机制。具体来说，我们将开发工具来（1）生成 具有复杂形状的可及膜，用于曲率依赖性蛋白质的定量研究， 膜相互作用，（2）构建具有动态可调张力的脂质体和脂质纳米盘，以研究 膜张力在调节蛋白质构象和脂质转运中的作用，以及（3）产生跨膜 具有可调尺寸和化学选择性的纳米孔，用于重构细胞器样隔室。我们 我希望这些新工具能够为结构生物学的研究提供支持，并具有潜在的变革性， 生物物理学、机械生物学和合成生物学。

PROJECT SUMMARY Lipid-bilayer membranes form diverse and dynamic structures in cells to serve vital functions including nutrient uptake, waste managements, signal transduction, and so on. Understanding the molecular mechanisms by which the cell generates and changes its membrane structures has been a central task of cell biology. It is well established that the physical and chemical properties of membranes regulate their interactions with proteins, which in turn shape the membrane landscape. However, there are still major knowledge gaps regarding how proteins act upon different membrane curvature and tension, especially when the membrane structure and/or the membrane-protein interaction are transient. Cell-free systems using reconstituted membranes provide a powerful method to study such intricate molecular interplays. However, there is still a pressing need for a precisely engineered platform that (1) exquisitely controls the geometrical, biochemical and mechanical properties of membranes and (2) easily allows for biochemical and structural characterization of membrane- associating proteins. Using DNA nanotechnology, a bottom-up method that generates three-dimensional molecular assemblies with programmable shape, stiffness, chemical modification, and motion, we propose to build nanoengineered membranes to bridge the technical gap in membrane manipulation and to unravel the mechanisms of protein-mediated membrane dynamics. Specifically, we will develop tools to (1) generate accessible membranes with complex shapes for the quantitative study of curvature-dependent protein- membrane interactions, (2) build liposomes and lipid nanodiscs with dynamically tunable tension to study the role of membrane tension in regulating protein conformation and lipid transport, and (3) create transmembrane nanopores with tunable size and chemical selectivity for the reconstitution of organelle-like compartments. We expect these new tools to be enabling and potentially transformative for research in structural biology, biophysics, mechanobiology, and synthetic biology.