The Structural Role of Kinesin Neck Region

运动蛋白，一种微管的运动蛋白，可以在微管上移动超过1。这可以通过一个模型来解释，即双头驱动蛋白分子的每个头可以交替地与微管相互作用。在这种情况下，每个头部之间必须有一定的协调，研究运动进程性的起源对于了解运动蛋白的分子机制是重要的。在这项研究中，我们使用光学镊子在纳米检测系统中测量了驱动蛋白，几种驱动蛋白-1样蛋白以及它们之间的嵌合体蛋白的加工能力。酪蛋白二聚体、酪蛋白单体、酪蛋白二聚体和酪蛋白单体在大肠杆菌中表达并纯化。这些蛋白质通过生物素-亲和素系统与微球结合，在没有ATP的情况下与微管结合的微球部分和在微管上运动超过20纳米的微球部分被评分。结果表明，ncd和kid的单个分子不能移动超过20 nm。我们进一步检测了嵌合体蛋白在kinesin和kid之间的运动性，发现具有kinesin头部的嵌合体蛋白单分子可以进行运动，而具有kinesin颈部的双头kid的嵌合体蛋白单分子不能进行运动。这些结果表明，两个运动蛋白头通过运动蛋白颈之间的协调并不是加工能力所必需的。

Kinesin, a motor protein for microtubules, can move on a microtubule processively more than I um. This is explained by a model that the each heads of a double-headed kinesin molecule can interact with a microtubule alternately. In that case, there must be some coordination between each heads, and it is important to investigate the origin of processivity of the movement to know the molecular mechanism of motor proteins. In this study, we measured the processivity of kinesin, several kinesin-1ike proteins, and chimera proteins between them in a nanometer detecting in vitro system using optical tweezers.Kinesin dimers, kinesin monomers, ncd dimers, and kid monomers were expressed in E. coli and then purified. These proteins were bound to microbeads via biotin-avidin system and the fractions of beads which show the binding to a microtubule in the absence of ATP and the fractions of beads which show the movement on a microtubule more than 20 nm were scored. The results showed that a single molecule of ncd and kid cannot move more than 20 nm. We further examined the motility of chimera proteins between kinesin and kid, and found that a single molecule of the chimera protein which has kinesin head can move processively, but a single molecule of the chimera protein which has double-headed kid via kinesin neck cannot move processively. These results suggest that the coordination between two kinesin heads via kinesin neck is not essential for processivity.