A universal pathway for kinesin stepping.

A universal pathway for kinesin stepping.
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DOI:
10.1038/nsmb.2104
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发表时间:
2011-08-14
影响因子:
16.8
通讯作者:
--
中科院分区:
生物学1区
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--
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驱动蛋白-1是一种ATP驱动的进行性马达,其在严格调节的步进循环中沿着微管运输货物。有效的门控机制确保动力学事件的顺序以适当的顺序进行,从而产生大量的连续反应循环。为了研究门控,我们创建了两个具有扩展颈连接器的突变体构建体,并使用单分子光捕获和系综荧光技术测量了它们的性质。由于头间张力的降低,构建体进入另外很少填充的构象状态,其中两个马达头保持与微管结合。在中等阻碍负荷下,观察到ATP依赖性、进行性反步和无效水解。基于测量,我们制定了一个全面的模型驱动蛋白运动,包括反应途径向前和向后步进。除了头间张力,我们发现,颈部连接器的方向也负责确保门控驱动蛋白。
Kinesin-1 is an ATP-driven, processive motor that transports cargo along microtubules in a tightly regulated stepping cycle. Efficient gating mechanisms ensure that the sequence of kinetic events proceeds in proper order, generating a large number of successive reaction cycles. To study gating, we created two mutant constructs with extended neck-linkers and measured their properties using single-molecule optical trapping and ensemble fluorescence techniques. Due to a reduction in the inter-head tension, the constructs access an otherwise rarely populated conformational state where both motor heads remain bound to the microtubule. ATP-dependent, processive backstepping and futile hydrolysis were observed under moderate hindering loads. Based on measurements, we formulated a comprehensive model for kinesin motion that incorporates reaction pathways for both forward and backward stepping. In addition to inter-head tension, we find that neck-linker orientation is also responsible for ensuring gating in kinesin.
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