The Mechanism of Kinesin Self-Regulation

驱动蛋白自我调节机制

• 批准号: 7487743
• 负责人: Sarah E. Rice
• 金额: $ 24.7万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-09-01 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7487743
• 关键词: A-Microtubule; ATP Hydrolysis; ATP phosphohydrolase; Address; Affect; Binding; Binding Sites; Charge; Electron Spin Resonance Spectroscopy; Elements; Frequencies; Head; In Vitro; Indium; Informal Social Control; Kinesin; Lead; Length; Light; Methods; Microtubules; Molecular Conformation; Movement; Nucleotides; Personal Satisfaction; Phosphorylation; Regulation; Research Personnel; Rice; Role; Tail; Testing; Tubulin; Work; dimer; drug discovery; in vivo; prevent; research study

DESCRIPTION (provided by applicant): The mechanism by which truncated kinesin dimers hydrolyze ATP and move 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 activation. 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.

描述（由申请人提供）：截短的驱动蛋白二聚体水解ATP并沿沿着单向移动的机制已被充分理解。目前还不清楚完整的驱动蛋白异源四聚体，其中有两个重链和两个轻链，是如何调节和激活的货物运输。在这项工作中，我们将测试的假设，驱动蛋白的调节时，尾巴直接绑定的头部，以防止ADP释放或微管结合。驱动蛋白可以通过其轻链和微管之间的电荷冲突进一步调节，并且当磷酸化轻链竞争尾部远离头部时，驱动蛋白可以被重新激活。我们将在四个具体目标中检验这些假设。前两个目的使用全长驱动蛋白重链解决调节，后两个目的探索轻链在调节和激活中的作用。在目标#1中，我们将确定尾部是否直接结合在微管结合位点或头部的核苷酸敏感元件上，或者它是否变构影响头部的核苷酸或微管结合区域。目标#2的实验由目标#1的结果指导，将确定头部的哪个区域结合尾部，并将鉴定特定的头部-尾部相互作用。在体内和体外进行的实验表明，轻链可能在调节驱动蛋白中具有重要作用，这将在目标#3中进行评估。最后，我们将确定驱动蛋白轻链的磷酸化是否可以直接激活目标#4中的驱动蛋白。总之，这些实验将扩大我们的相互作用和构象变化，支配驱动蛋白活动的理解。此外，在这项工作中发现的调节相互作用可能揭示了在几种驱动蛋白中相似的抑制机制。这可能会导致更快地发现专门针对驱动蛋白的药物。