Kluger Carleen, Braun Lukas, Sedlak Steffen M, Pippig Diana A, Bauer Magnus S, Miller Ken, Milles Lukas F, Gaub Hermann E, Vogel Viola
Lehrstuhl für Angewandte Physik and Center for NanoScience, Ludwig-Maximilians-Universität München, Munich, Germany.
Laboratory of Applied Mechanobiology, Department of Health Sciences and Technology, ETH Zurich, Zurich, Switzerland.
Biophys J. 2020 Mar 24;118(6):1344-1356. doi: 10.1016/j.bpj.2019.12.042. Epub 2020 Feb 4.
Vinculin is a universal adaptor protein that transiently reinforces the mechanical stability of adhesion complexes. It stabilizes mechanical connections that cells establish between the actomyosin cytoskeleton and the extracellular matrix via integrins or to neighboring cells via cadherins, yet little is known regarding its mechanical design. Vinculin binding sites (VBSs) from different nonhomologous actin-binding proteins use conserved helical motifs to associate with the vinculin head domain. We studied the mechanical stability of such complexes by pulling VBS peptides derived from talin, α-actinin, and Shigella IpaA out of the vinculin head domain. Experimental data from atomic force microscopy single-molecule force spectroscopy and steered molecular dynamics (SMD) simulations both revealed greater mechanical stability of the complex for shear-like than for zipper-like pulling configurations. This suggests that reinforcement occurs along preferential force directions, thus stabilizing those cytoskeletal filament architectures that result in shear-like pulling geometries. Large force-induced conformational changes in the vinculin head domain, as well as protein-specific fine-tuning of the VBS sequence, including sequence inversion, allow for an even more nuanced force response.
纽蛋白是一种普遍存在的衔接蛋白，可短暂增强黏附复合物的机械稳定性。它稳定了细胞通过整合素在肌动球蛋白细胞骨架与细胞外基质之间或通过钙黏着蛋白与相邻细胞之间建立的机械连接，但对其机械设计却知之甚少。来自不同非同源肌动蛋白结合蛋白的纽蛋白结合位点（VBS）利用保守的螺旋基序与纽蛋白头部结构域结合。我们通过将源自踝蛋白、α-辅肌动蛋白和志贺氏菌IpaA的VBS肽从纽蛋白头部结构域中拉出，研究了此类复合物的机械稳定性。原子力显微镜单分子力谱和定向分子动力学（SMD）模拟的实验数据均表明，与拉链状拉伸构型相比，复合物在剪切状拉伸构型下具有更高的机械稳定性。这表明增强作用沿着优先的力方向发生，从而稳定了那些导致剪切状拉伸几何形状的细胞骨架丝结构。纽蛋白头部结构域中由力诱导的大的构象变化，以及包括序列反转在内的VBS序列的蛋白质特异性微调，允许产生更细微的力响应。
