Shrestha Prakash, Yang Darren, Shih William M, Wong Wesley P
bioRxiv. 2023 Jul 11:2023.07.10.548386. doi: 10.1101/2023.07.10.548386.
The ability to accurately map the 3D geometry of single-molecule complexes in trace samples would lead to new insights into molecular mechanics and provide an approach for single-molecule structural proteomics. To enable this, we have developed a high-resolution force-spectroscopy method capable of measuring multiple distances between labeled sites in natively folded protein complexes. Our approach combines reconfigurable nanoscale devices we call DNA Nanoswitch Calipers, which we have previously introduced, with a force-based barcoding system to distinguish each measurement location. We demonstrate our approach by reconstructing the tetrahedral geometry of biotin-binding sites in natively folded streptavidin, with 1.5-2.5 agreement to previously reported structures.
精确绘制痕量样品中单分子复合物的三维几何结构，这一能力将为分子力学带来新的见解，并为单分子结构蛋白质组学提供一种方法。为实现这一目标，我们开发了一种高分辨率力谱方法，该方法能够测量天然折叠的蛋白质复合物中标记位点之间的多个距离。我们的方法将我们之前介绍过的称为DNA纳米开关卡尺的可重构纳米级装置与基于力的条形码系统相结合，以区分每个测量位置。我们通过重建天然折叠的链霉亲和素中生物素结合位点的四面体几何结构来证明我们的方法，与先前报道的结构的一致性为1.5 - 2.5 。
