Cellular and Molecular Biology Program, University of Michigan, Ann Arbor, United States.
Department of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, United States.
Elife. 2019 May 14;8:e44146. doi: 10.7554/eLife.44146.
Kinesin force generation involves ATP-induced docking of the neck linker (NL) along the motor core. However, the roles of the proposed steps of NL docking, cover-neck bundle (CNB) and asparagine latch (N-latch) formation, during force generation are unclear. Furthermore, the necessity of NL docking for transport of membrane-bound cargo in cells has not been tested. We generated kinesin-1 motors impaired in CNB and/or N-latch formation based on molecular dynamics simulations. The mutant motors displayed reduced force output and inability to stall in optical trap assays but exhibited increased speeds, run lengths, and landing rates under unloaded conditions. NL docking thus enhances force production but at a cost to speed and processivity. In cells, teams of mutant motors were hindered in their ability to drive transport of Golgi elements (high-load cargo) but not peroxisomes (low-load cargo). These results demonstrate that the NL serves as a mechanical element for kinesin-1 transport under physiological conditions.
驱动蛋白力的产生涉及到 ATP 诱导的颈部接头（NL）沿着马达核心的对接。然而，NL 对接、覆盖颈束（CNB）和天冬酰胺闩锁（N-闩锁）形成的提议步骤在力的产生过程中的作用尚不清楚。此外，NL 对接对于细胞内膜结合货物运输的必要性尚未得到测试。我们基于分子动力学模拟生成了在 CNB 和/或 N-闩锁形成方面受损的驱动蛋白-1 马达。突变体马达在光阱测定中显示出降低的力输出和无法失速的能力，但在空载条件下表现出增加的速度、运行长度和着陆率。因此，NL 对接虽然提高了力的产生，但却以速度和连续性为代价。在细胞中，突变体马达在驱动高尔基体元件（高负荷货物）运输方面的能力受到阻碍，但不能驱动过氧化物酶体（低负荷货物）运输。这些结果表明，NL 作为一种机械元件，在生理条件下为驱动蛋白-1 的运输提供服务。
