Department of Physics, University of Osnabrück, Osnabrück, Germany.
Biophys J. 2011 Oct 5;101(7):1772-80. doi: 10.1016/j.bpj.2011.08.048.
The cytoskeletal adaptor protein vinculin plays an important role in the control of cell adhesion and migration, linking the actin cytoskeleton to adhesion receptor complexes in cell adhesion sites. The conformation of the vinculin tail dimer, which is crucial for protein function, was analyzed using site-directed spin labeling in electron paramagnetic resonance spectroscopy. Interspin distances for a set of six singly and four doubly spin-labeled mutants of the tail domain of vinculin were determined and used as constraints for modeling of the vinculin tail dimer. A comparison of the results obtained by molecular dynamic simulations and a rotamer library approach reveals that the crystal structure of the vinculin tail monomer is essentially preserved in aqueous solution. The orientation of monomers within the dimer observed previously by x-ray crystallography agrees with the solution electron paramagnetic resonance data. Furthermore, the distance between positions 1033 is shown to increase by >3 nm upon interaction of the vinculin tail domain with F-actin.
细胞骨架衔接蛋白 vinculin 在控制细胞黏附和迁移方面发挥着重要作用，它将肌动蛋白细胞骨架与细胞黏附部位的黏附受体复合物连接起来。使用电子顺磁共振波谱中的定点自旋标记技术分析了 vinculin 尾部二聚体的构象，该构象对蛋白质功能至关重要。确定了一组六个单自旋标记和四个双自旋标记突变体的自旋间距离，并将其用作 vinculin 尾部二聚体建模的约束条件。通过分子动力学模拟和构象文库方法的比较，结果表明 vinculin 尾部单体的晶体结构在水溶液中基本保持不变。先前通过 X 射线晶体学观察到的二聚体中单体的取向与溶液电子顺磁共振数据一致。此外，研究表明，当 vinculin 尾部结构域与 F-肌动蛋白相互作用时，位置 1033 之间的距离增加了>3nm。
