MINFLUX dissects the unimpeded walking of kinesin-1
MINFLUX dissects the unimpeded walking of kinesin-1
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DOI：
10.1126/science.ade2650
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发表时间：
2023-03-10
期刊：
影响因子：
56.9
通讯作者：
Hell, Stefan W.
中科院分区：
文献类型：
--
作者：
Wolff, Jan O.;Scheiderer, Lukas;Hell, Stefan W.
我们介绍了一种干涉MINFLUX显微镜，记录蛋白质运动高达1.7纳米每毫秒的时空精度。这种精确度以前需要将不成比例的大珠子连接到蛋白质上，但MINFLUX只需要从大约1纳米大小的荧光团中检测大约20个光子。因此，我们能够研究运动蛋白驱动蛋白-1在高达生理三磷酸腺苷（ATP）浓度的微管上的步进。我们发现旋转的柄和无负荷驱动蛋白的头部在步进，并表明，ATP是采取了一个单一的头部绑定到微管和ATP水解发生时，两个头部都绑定。我们的研究结果表明，MINFLUX量化（亚）毫秒的蛋白质构象变化与最小的干扰。
We introduce an interferometric MINFLUX microscope that records protein movements with up to 1.7 nanometer per millisecond spatiotemporal precision. Such precision has previously required attaching disproportionately large beads to the protein, but MINFLUX requires the detection of only about 20 photons from an approximately 1-nanometer-sized fluorophore. Therefore, we were able to study the stepping of the motor protein kinesin-1 on microtubules at up to physiological adenosine-5 ' triphosphate (ATP) concentrations. We uncovered rotations of the stalk and the heads of load-free kinesin during stepping and showed that ATP is taken up with a single head bound to the microtubule and that ATP hydrolysis occurs when both heads are bound. Our results show that MINFLUX quantifies (sub)millisecond conformational changes of proteins with minimal disturbance.