BUDDING YEAST SEPTIN FILAMENTS: CONFORMATION AND KINETICS OF FILAMENT FORMATION

出芽酵母败丝：丝形成的构象和动力学

• 批准号: 8170202
• 负责人: AURELIE BERTIN
• 金额: $ 0.17万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8170202
• 关键词: Cell division; Cells; Complex; Computer Retrieval of Information on Scientific Projects Database; Cytokinesis; Cytoskeleton; Electron Microscopy; Filament; Funding; Grant; In Vitro; Institution; Ionic Strengths; Kinetics; Label; Mitotic; Molecular; Molecular Conformation; Mothers; Neck; Nucleotides; Polymers; Process; Proteins; Research; Research Personnel; Resources; Saccharomyces cerevisiae; Saccharomycetales; Sodium Chloride; Source; Techniques; Testing; United States National Institutes of Health; Yeasts; daughter cell; flexibility; mutant; polymerization; research study

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. Septin proteins assemble into a cytoskeletal polymer. Septin cytoskeleton is essential for cytokinesis, the last step in cell division. In budding yeast, filamentous rings of septins have been visualized at the bud neck and constrict it in order to separate the mother and daughter cells. We are using yeast septins that self-assemble in vitro. Mitotic Saccharomyces cerevisiae cells express five septins: Cdc3, Cdc10, Cdc11, Cdc12, and Shs1/Sep7. All, but Shs1, are essential for cell division. We have determined the molecular organization of the four essential septins within the minimal complex by labeling techniques using electron microscopy. All of the septins but Cdc10 have a predicted coiled coil which promotes septin assembly into paired filaments. Octameric minimal subunits assemble into filaments when the ionic strength is lowered to 200 mM monovalent salt. Small angle x-ray scattering will be used on both wild type septins and on mutant septins deprived of coiled coils to be able to locate those flexible domains. In addition to that, using a stopped flow mixer, the kinetics of filament formation will be probed. Those kinetics experiments will be performed in the presence of various nucleotides so their effect on the polymerization process can be tested.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目和 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 Septin 蛋白组装成细胞骨架聚合物。 Septin 细胞骨架对于胞质分裂（细胞分裂的最后一步）至关重要。在芽殖酵母中，在芽颈处可以看到丝状的脓蛋白环，并将其收缩以分离母细胞和子细胞。我们使用的是在体外自组装的酵母败血症。有丝分裂的酿酒酵母细胞表达五种脓蛋白：Cdc3、Cdc10、Cdc11、Cdc12 和 Shs1/Sep7。除 Shs1 外，所有这些对于细胞分裂都是必需的。我们通过使用电子显微镜的标记技术确定了最小复合物内四种基本脓毒症的分子组织。除 Cdc10 之外的所有脓蛋白都具有预测的卷曲线圈，可促进脓蛋白组装成成对的细丝。当离子强度降低至 200 mM 一价盐时，八聚体最小亚基组装成丝。小角度 X 射线散射将用于野生型脓毒症和缺乏卷曲线圈的突变型脓毒症，以便能够定位这些柔性结构域。除此之外，使用停流混合器，将探测细丝形成的动力学。这些动力学实验将在各种核苷酸存在的情况下进行，以便测试它们对聚合过程的影响。