Distinct Interaction Modes of the Kinesin-13 Motor Domain with the Microtubule
Distinct Interaction Modes of the Kinesin-13 Motor Domain with the Microtubule
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DOI：
10.1016/j.bpj.2016.02.029
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发表时间：
2016-04-12
影响因子：
3.4
通讯作者：
Sosa, Hernando
中科院分区：
文献类型：
--
作者：
Chatterjee, Chandrima;Benoit, Matthieu P. M. H.;Sosa, Hernando
驱动蛋白-13是运动蛋白的驱动蛋白超家族的成员，其使微管（MT）解聚并且没有运动活性。与大多数其他驱动蛋白一样，驱动蛋白-13在MT晶格上产生单向运动，而不是进行一维扩散（ODD）并在MT末端诱导解聚。为了了解ODD的机制和不同的驱动蛋白-13功能的起源，我们使用了合奏和单分子荧光偏振显微镜分析的行为和构象的果蝇黑腹驱动蛋白-13 KLP 10A蛋白质结构绑定到MT晶格。我们发现KLP 10 A与MT以两种共存模式相互作用：一种是马达结构域以特定方向结合到MT晶格，另一种是马达结构域非常移动的并且能够进行ODD。通过比较突变和缺失构建体的取向和动力学行为，我们得出结论：1）驱动蛋白-13类特异性颈结构域和环-2有助于相对于MT定向运动结构域。2)在ODD期间，KLP 10A马达域迅速改变方向（岩石或翻滚）。3)运动域单独能够经历ODD。4)KLP 10A运动域中的第二个微管蛋白结合位点对ODD并不重要。5)颈部结构域不是阻止KLP 10A与MT晶格结合的元件，如运动驱动蛋白。
Kinesins-13s are members of the kinesin superfamily of motor proteins that depolymerize microtubules (MTs) and have no motile activity. Instead of generating unidirectional movement over the MT lattice, like most other kinesins, kinesins-13s undergo one-dimensional diffusion (ODD) and induce depolymerization at the MT ends. To understand the mechanism of ODD and the origin of the distinct kinesin-13 functionality, we used ensemble and single-molecule fluorescence polarization microscopy to analyze the behavior and conformation of Drosophila melanogaster kinesin-13 KLP10A protein constructs bound to the MT lattice. We found that KLP10A interacts with the MT in two coexisting modes: one in which the motor domain binds with a specific orientation to the MT lattice and another where the motor domain is very mobile and able to undergo ODD. By comparing the orientation and dynamic behavior of mutated and deletion constructs we conclude that 1) the Kinesin-13 class specific neck domain and loop-2 help orienting the motor domain relative to the MT. 2) During ODD the KLP10A motor-domain changes orientation rapidly (rocks or tumbles). 3) The motor domain alone is capable of undergoing ODD. 4) A second tubulin binding site in the KLP10A motor domain is not critical for ODD. 5) The neck domain is not the element preventing KLP10A from binding to the MT lattice like motile kinesins.