The Mechanism of Kinesin Self-Regulation

截短的动蛋白二聚体水解三磷酸腺苷并沿其单向运动的机制 人们对微管有很好的了解。目前还不太清楚全动能如何在异构体中存在，它有两个重的 链条和两条轻链被管制和激活，用于货物运输。在这项工作中，我们将测试 当尾巴直接捆绑头部以防止ADP释放或 微管结合。Kinesin的轻链和轻链之间的电荷冲突可能会进一步调节它 当磷酸化轻链与尾巴竞争时，微管和动蛋白可能会重新激活 人头。我们将在四个具体目标上检验这些假设。前两个目标是通过使用 全长运动在重链中，后两个目的是探索轻链在调节中的作用 和激活度。在目标1中，我们将确定尾巴是直接结合在微管结合部位还是与微管结合部位结合 头部的核苷酸感应元件，或者它是否变构地影响核苷酸-或者 头部微管结合区。在目标1的结果指导下，目标2的实验将 确定头部的哪个区域绑定尾巴，并将识别特定的头尾相互作用。实验 在体内和体外进行的研究表明，轻链可能在调节 激动素，这将在目标3中进行评估。最后，我们将确定光 链可以直接激活Aim#4中的Kinesin。这些实验将扩展我们对 控制动蛋白活性的相互作用和构象变化。此外，监管机构 在这项工作中发现的相互作用可能揭示了几种动蛋白相似的抑制机制。 这可能会导致更快地发现专门针对肌动蛋白的药物。

The mechanism by which truncated kinesin dimers hydrolyze ATP and moves unidirectionally along microtubules is well understood. It is far less clear how the full kinesin heterotetramer, which has two heavy chains and two light chains, is regulated and activated for cargo transport. In this work, we will test the hypothesis that kinesin is regulated when the tails directly bind the heads to prevent ADP release or microtubule binding. Kinesin may be further regulated by a charge clash between its light chains and microtubules, and kinesin may be re-activated when phosphorylated light chains compete the tails away from the heads. We will test these hypotheses in four Specific Aims. The first two Aims address regulation using the full-length kinesin heavy chain, and the second two Aims explore the role of the light chains in regulation and activiation. In Aim #1, we will determine whether the tail binds directly in the microtuble-binding site or to the nucleotide-sensing elements in the head, or whether it allosterically affects the nucleotide- or microtubule-binding regions of the head. The experiments of Aim #2, guided by the results of Aim #1, will determine what region of the head binds the tail and will identify specific head-tail interactions. Experiments performed both in vivo and in vitro indicate that the light chains may have a significant role in regulating kinesin, which will be assessed in Aim #3. Lastly, we will determine whether phosphorylation of kinesin light chains can directly activate kinesin in Aim #4. Together, these experiments will extend our understanding of the interactions and conformational changes that govern kinesin activity. Furthermore, the regulatory interactions that are found in this work may reveal inhibitory mechanisms that are similar in several kinesins. This may lead to quicker discovery of drugs that specifically target kinesins.