Single Molecule Translocation through Biological Nanopores with Optical Tweezers

在该研究项目中，我们计划使用光镊子力力学以（亚）分子分辨率研究和量化单个生物聚合物通过生物纳米孔（生物孔）的转运机制和动力学。这种单分子技术允许在没有分子标记和扩增的情况下测量有效的分子力和弹性、机械噪声谱、易位动力学、相关的能量景观以及相关的纳米流体特性。我们将研究链状大分子，如核酸、多糖以及折叠和未折叠的蛋白质如何穿过生物孔（孔），这些孔（孔）在功能上集成在支持的生物膜和仿生脂质混合物中，将单分子力光谱的多个方面与纳米流体相结合。由于单分子易位受热非平衡、快速热波动和大分子随机性的控制，我们的目标是进行快速力和噪声分析，并通过随机模拟对实验结果进行一致描述（合作：P. Reimann）。我们的目标是深入了解通过生物孔进行分子易位的基本机制，以及纳米生物技术的应用。后者依赖于通过生物纳米孔在易位过程中识别特定的亚分子力特征，从而推断易位大分子的结构信息（例如DNA的甲基化模式）。

In the proposed research project we plan to use optical tweezers force mechanics to investigate and quantify the translocation mechanisms and dynamics of single biopolymers through biological nanopores (biopores) with (sub)molecular resolution. This single molecule technology allows to measure effective molecular forces and elasticities, mechanical noise spectra, translocation dynamics, associated energy landscape as well as associated nanofluidic properties without molecule labelling and amplification. We will study how chain-like macromolecules like nucleic acids, polysaccharides, as well as folded and unfolded proteins will be threaded through biopores (porins) that are functionally integrated in supported biomembranes and biomimetic lipid-hybrids thereof, combining manifold aspects of single-molecule force spectroscopy with nanofluidics. Since single-molecule translocation is governed by thermal non-equilibrium, fast thermal fluctuations and macromolecular stochastics, we aim for a fast force and noise analysis together with a coherent description of the experimental results with stochastic simulations (Cooperation: P. Reimann). We aim for deep fundamental mechanistic insights into the molecule translocation through biopores in general, as well as nanobiotechnological applications thereof. The latter rely on the identification of specific, sub-molecular force signatures during translocation through biological nanopores, allowing to deduce structural information of the translocated macromolecule (e.g. the methylation pattern of DNA).