N‐terminal β‐strand of single‐headed kinesin‐1 can modulate the off‐axis force‐generation and resultant rotation pitch

N‐terminal β‐strand of single‐headed kinesin‐1 can modulate the off‐axis force‐generation and resultant rotation pitch
复制标题

单头驱动蛋白-1的N端β链可以调节离轴力的产生和由此产生的旋转螺距

DOI:
10.1002/cm.21630
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发表时间:
2020
期刊:
影响因子:
2.9
通讯作者:
Yajima Junichiro
Yajima Junichiro
中科院分区:
生物学4区
文献类型:
--
作者:
Yamagishi Masahiko;Fujimura Shoko;Sugawa Mitsuhiro;Nishizaka Takayuki;Yajima Junichiro

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在体外微管滑动测定中,大多数驱动蛋白驱动滑动微管以螺旋运动绕其纵轴旋转。螺旋螺距小于微管的超扭曲原丝螺距,表明螺旋螺距是驱动蛋白的固有特性。为了阐明驱动蛋白螺旋微管的分子机制,我们在各种测定条件下对与微管结合的量子点进行了三维跟踪,该量子点在涂有单头驱动蛋白-1的表面上易位,以改变驱动蛋白和微管之间的相互作用。尽管驱动蛋白浓度、离子强度和 ATP 浓度的变化改变了滑动和旋转速度,但在所有测试条件下,除了在低 ATP 浓度下螺距略有增加外,螺旋螺距仍保持左旋且恒定在约 0.3 μm。然后,我们使用我们的系统分析了从驱动蛋白运动核心突出的 N 端 β 链点突变的影响,发现了减少螺旋螺距的突变。我们的研究结果证实,微管的螺旋运动是由驱动蛋白的内在特性引起的,并表明源自头部 N 端 β 链的主动力或阻滞力的变化可调节音调。
In in vitro microtubule gliding assays, most kinesins drive the rotation of gliding microtubules around their longitudinal axes in a corkscrew motion. The corkscrewing pitch is smaller than the supertwisted protofilament pitch of microtubules, indicating that the corkscrewing pitch is an inherent property of kinesins. To elucidate the molecular mechanisms through which kinesins corkscrew the microtubule, we performed three‐dimensional tracking of a quantum dot bound to a microtubule translocating over a surface coated with single‐headed kinesin‐1 s under various assay conditions to alter the interactions between the kinesin and microtubule. Although alternations in kinesin concentration, ionic strength, and ATP concentration changed both gliding and rotational velocities, the corkscrewing pitch remained left‐handed and constant at ~0.3 μm under all tested conditions apart from a slight increase in pitch at a low ATP concentration. We then used our system to analyze the effect of point mutations in the N‐terminal β‐strand protruding from the kinesin motor core and found mutations that decreased the corkscrewing pitch. Our findings confirmed that the corkscrewing motion of microtubules is caused by the intrinsic properties of the kinesin and demonstrates that changes in the active or retarding force originating from the N‐terminal β‐strand in the head modulate the pitch.
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