Department of Chemistry and Biochemistry, University of Delaware, Newark, DE, USA.
Department of Molecular Medicine, Beckman Research Institute of City of Hope, Duarte, CA, USA.
Nat Commun. 2022 Nov 10;13(1):6795. doi: 10.1038/s41467-022-34026-w.
Microtubules (MTs) and their associated proteins play essential roles in maintaining cell structure, organelle transport, cell motility, and cell division. Two motors, kinesin and cytoplasmic dynein link the MT network to transported cargos using ATP for force generation. Here, we report an all-atom NMR structure of nucleotide-free kinesin-1 motor domain (apo-KIF5B) in complex with paclitaxel-stabilized microtubules using magic-angle-spinning (MAS) NMR spectroscopy. The structure reveals the position and orientation of the functionally important neck linker and how ADP induces structural and dynamic changes that ensue in the neck linker. These results demonstrate that the neck linker is in the undocked conformation and oriented in the direction opposite to the KIF5B movement. Chemical shift perturbations and intensity changes indicate that a significant portion of ADP-KIF5B is in the neck linker docked state. This study also highlights the unique capability of MAS NMR to provide atomic-level information on dynamic regions of biological assemblies.
微管（MTs）及其相关蛋白在维持细胞结构、细胞器运输、细胞运动和细胞分裂方面发挥着重要作用。两种马达蛋白，驱动蛋白和细胞质动力蛋白，利用 ATP 为产生力将 MT 网络与运输货物连接起来。在这里，我们使用魔角旋转（MAS）NMR 光谱法报道了无核苷酸的驱动蛋白-1 马达结构域（apo-KIF5B）与紫杉醇稳定的微管复合物的全原子 NMR 结构。该结构揭示了功能重要的颈环接头的位置和取向，以及 ADP 如何诱导颈环接头随之发生的结构和动力学变化。这些结果表明，颈环接头处于未对接构象，并且定向与 KIF5B 运动的方向相反。化学位移扰动和强度变化表明，相当一部分 ADP-KIF5B 处于颈环接头对接状态。这项研究还强调了 MAS NMR 提供生物组装体动态区域原子水平信息的独特能力。
