The Structural Basis of Force Generation by the Mitotic Motor Kinesin-5
The Structural Basis of Force Generation by the Mitotic Motor Kinesin-5
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DOI：
10.1074/jbc.m112.404228
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发表时间：
2012-12-28
影响因子：
4.8
通讯作者：
Moores, Carolyn A.
中科院分区：
文献类型：
--
作者：
Goulet, Adeline;Behnke-Parks, William M.;Moores, Carolyn A.
在有丝分裂过程中，需要Kinesin-5来形成双极纺锤体。它的运动域包含核苷酸和微管结合位点以及产生力量的机械元件，已经为其基于纺锤体的功能进化出了不同的特性。在这项研究中，我们报道了核苷酸结合前后微管结合的人Kinesin-5的亚纳米分辨率冷冻电子显微镜重建，并将这些信息与核苷酸诱导的颈链和覆盖链移动的动力学研究相结合。这些研究揭示了依赖核苷酸的偶联构象变化，解释了这种马达的许多特性。我们发现，ATP结合诱导颈部连接子的棘轮状对接和N-末端覆盖链的同时平行对接。当ATP结合时，变构抑制剂Kinesin-5的结合部位L5环也经历了戏剧性的重新定位，表明它直接参与了核苷酸结合的控制。我们的结构表明，人类动蛋白-5的变构抑制剂正被开发为抗癌治疗药物，它与正常功能过程中发生的运动构象结合。然而，由于L5中进化定义的序列变异，无脊椎动物Kinesin-5s不采用这种构象，这解释了它们对药物抑制的抗性。总而言之，我们的数据揭示了Kinesin-5马达的分子机制进化到力产生的精确度。
Kinesin-5 is required for forming the bipolar spindle during mitosis. Its motor domain, which contains nucleotide and microtubule binding sites and mechanical elements to generate force, has evolved distinct properties for its spindle-based functions. In this study, we report subnanometer resolution cryoelectron microscopy reconstructions of microtubule-bound human kinesin-5 before and after nucleotide binding and combine this information with studies of the kinetics of nucleotide-induced neck linker and cover strand movement. These studies reveal coupled, nucleotide-dependent conformational changes that explain many of this motor's properties. We find that ATP binding induces a ratchet-like docking of the neck linker and simultaneous, parallel docking of the N-terminal cover strand. Loop L5, the binding site for allosteric inhibitors of kinesin-5, also undergoes a dramatic reorientation when ATP binds, suggesting that it is directly involved in controlling nucleotide binding. Our structures indicate that allosteric inhibitors of human kinesin-5, which are being developed as anti-cancer therapeutics, bind to a motor conformation that occurs in the course of normal function. However, due to evolutionarily defined sequence variations in L5, this conformation is not adopted by invertebrate kinesin-5s, explaining their resistance to drug inhibition. Together, our data reveal the precision with which the molecular mechanism of kinesin-5 motors has evolved for force generation.