TOPOISOMERASES

拓扑异构酶

• 批准号: 7598612
• 负责人: LYNN ZECHIEDRICH
• 金额: $ 1.63万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-12-01 至 2007-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7598612
• 关键词: ATPase Domain; Antineoplastic Agents; Biochemical; Catenated DNA; Complex; Computer Retrieval of Information on Scientific Projects Database; Cryoelectron Microscopy; DNA; Data; Dimensions; Enzymes; Funding; Grant; Human; Individual; Institution; Kinetics; Maintenance; Metabolism; Neurofibrillary Tangles; Organism; Pharmacologic Substance; Proteins; Reaction; Research; Research Personnel; Resources; Source; Structure; Superhelical DNA; Topoisomerase; United States National Institutes of Health; Yeasts; enzyme structure; particle; reconstruction; three dimensional structure

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Topoisomerases are essential enzymes that impact nearly every aspect of DNA metabolism through the maintenance of DNA supercoiling, untangling DNA knots, and resolving DNA catenanes. These enzymes are highly conserved in all organisms and are major pharmaceutical targets. Our current challenges focus on how human topoisomerase IIalpha, a type-2 topoisomerase, untangles rather than tangles its DNA substrates. Although there are extensive biochemical data available for the enzyme, there exists no atomic structure information on any complete type-2 topoisomerase. Atomic structures are available of individual domains of eukaryotic type-2 enzymes, such as the ATPase domain for both the yeast and human enzymes and the cleavage-religation region of the yeast enzyme; however, there is no atomic structure information available for 22% of the C-terminus of the human protein. Electron cryomicroscopy and single particle reconstruction will be used to determine the three-dimensional structure of the complete enzyme alone or complexed with its DNA substrate in the presence and absence of anticancer drugs. In addition, reconstructions of isolated reaction intermediates will be used to determine the conformational changes of the enzyme. Human topoisomerase IIalpha is a homodimer of 340 kDa and our initial multiple refinement convergence reconstructions of the enzyme alone show the dimensions of the particles to be roughly 65 ¿ x 70 ¿ x 75 ¿. With the structures of the enzyme alone, complexed with DNA, with anticancer drugs, and each trapped kinetic intermediate, we will be able to interpret how the machine moves and progresses through its catalytic cycle.

该副本是使用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这是调查员的机构。 拓扑异构酶是必不可少的酶，通过维持DNA超串联，弄清DNA结和解决DNA Catenanes，几乎影响了DNA代谢的各个方面。这些酶在所有生物体中都是高度保守的，是主要的药物靶标。我们目前的挑战集中在人类拓扑异构酶iialpha（一种2型拓扑异构酶）如何解开而不是缠结其DNA底物。尽管有广泛的生化数据可用于酶，但没有关于任何完整类型2拓扑异构酶的原子结构信息。原子结构可用于真核2型酶的各个域，例如酵母和人类酶的ATPase结构域以及酵母酶的切割区域的裂解区域；但是，没有人类蛋白的C末端的22％的原子结构信息。 电子冷冻显微镜和单个颗粒重建将用于在存在和不存在抗癌药物的情况下单独确定完整酶的三维结构或与其DNA底物复合。此外，分离反应中间体的重建将用于确定酶的构象变化。人拓扑异构酶iiAlpha是340 kDa的同二聚体，我们最初的多个酶的多次完善收敛重建单独的重建表明，颗粒的尺寸大约为65€x 70€x 75€x 75？x 75€与单独使用的酶的结构与抗dna的结构相吻合，并与抗抗体构成的抗体，以及对抗抗药体的融合，我们的机器是如何的，我们的机器是抗抗菌药物的构造。通过其催化循环移动和进展。