SAXS STUDIES ON P1 PARTITION COMPLEXES

P1 划分复合体的 SAXS 研究

• 批准号: 7954359
• 负责人: Maria Schumacher
• 金额: $ 0.02万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-03-01 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7954359
• 关键词: ATP phosphohydrolase; Binding; Binding Sites; Boxing; Cell division; Centromere; Chromosome Positioning; Complex; Computer Retrieval of Information on Scientific Projects Database; DNA; DNA-Binding Proteins; E coli dnaG protein; Escherichia coli; Funding; Genetic Materials; Goals; Grant; Institution; Integration Host Factors; Mediating; Modeling; Movement; Plasmids; Pre-Par; Process; Proteins; Research; Research Personnel; Resolution; Resources; Role; Site; Source; Structure; System; Time; United States National Institutes of Health; Upper arm; Walkers; daughter cell; dimer; segregation; structural biology; synchrotron radiation

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. The faithful inheritance of prokaryotic genetic material requires the directed movement and positioning of chromosomes and plasmids to daughter cells at cell division. This process, called partition or segregation, is mediated by functionally homologous par systems comprised of a cis-acting centromere-like DNA site(s) and two proteins, ParA and ParB. The Escherichia coli P1 plasmid partition apparatus has served as a paradigm for partition. P1 ParA is a 44 kDa Walker type ATPase that drives plasmid separation at the final step of partition. P1 ParB is a 38 kDa DNA-binding protein that mediates the initial steps in segregation; partition complex formation and pairing. In partition complex formation, ParB and the E. coli protein, integration host factor (IHF), bind cooperatively to the ~74 bp parS centromere-like site, which contains multiple A- and B-Boxes, to form the partition complex. Intrinsically bent DNA can substitute for IHF, confirming that its role is simply to bring together the A-Box/B-Box containing parS arms, which bind ParB. After the initial partition complex is formed, ParB mediates pairing between plasmids as multiple ParB molecules load onto parS. Although it has been biochemically well characterized, a detailed mechanistic understanding of partition is lacking due, in large part, to the dearth of structural information on partition proteins and their complexes. Thus, the long terms goals of this proposal are to use the P1 par system as a model to study various steps in segregation. Our first goal is to obtain a low resolution structure of the initial ParB-IHF-parS partition complex. To do this we will utilize a parS site that allows binding of only a single ParB dimer. Subsequently, we will include additional binding sites to build larger partition complexes allowing us to view plasmid pairing for the first time and ultimately to trap a ParB-IHF-ParA(ATP)-parS pre-segregation complex.

该副本是利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，这不一定是调查员的机构。 实质性遗传物质的忠实遗传需要将染色体和质粒定向和定位给细胞分裂的子细胞。 该过程称为分区或隔离，是由由顺式作用的丝粒样DNA位点（S）和两个蛋白（PARA和PARB）组成的功能同源PAR系统介导的。 大肠杆菌P1质粒分区设备已成为分区的范式。 P1 Para是44 kDa Walker型ATPase，在分区的最后一步驱动质粒分离。 P1 PARB是一种38 kDa DNA结合蛋白，可介导隔离的初始步骤。分区复合物的形成和配对。 在分区复合物的形成中，PARB和大肠杆菌蛋白（Integnation host因子（IHF））与〜74 bp pars centromere样位点合作结合，其中包含多个A-和B盒形成分区复合物。 本质上弯曲的DNA可以代替IHF，证实其作用仅仅是将包含PARS的A-box/b-box组合在一起，该臂结合了PARB。 在形成初始分区复合物后，PARB将质粒之间的配对介导，因为多个PARB分子负载到PAR上。尽管它在生物化学上的特征很好，但在很大程度上缺乏对分区蛋白及其复合物的结构信息的详细机械理解。 因此，该提案的长期目标是将P1 PAR系统用作研究隔离的各种步骤的模型。 我们的第一个目标是获得初始PARB-IHF-PARS分区复合物的低分辨率结构。 为此，我们将使用一个PARS位点，该站点仅允许单个PARB二聚体绑定。 随后，我们将包括其他绑定位点，以构建较大的分区络合物，使我们能够首次查看质粒配对，并最终捕获PARB-IHF-PARA（ATP）PARS-PARS-PARS-PARS-PARS-PARS-PRE-pre-e-pre-egregegation Complect。