FISSION PROTEIN

裂变蛋白

• 批准号: 8168583
• 负责人: R Blake Hill
• 金额: $ 0.22万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-01-15 至 2010-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8168583
• 关键词: Adaptor Signaling Protein; Biochemical; C-terminal; Coiled-Coil Domain; Complement; Computer Retrieval of Information on Scientific Projects Database; Dynamin; Funding; Goals; Grant; Guanosine Triphosphate; Guanosine Triphosphate Phosphohydrolases; Health; Human; Hydrolysis; Image; Institution; Membrane; Mitochondria; Modeling; Molecular; N-terminal; Negative Staining; Process; Proteins; Recruitment Activity; Research; Research Personnel; Resources; Sampling; Site; Source; Structural Models; System; Tail; Testing; Transmembrane Domain; United States National Institutes of Health; Work; Yeasts; base; maltose-binding protein; particle; peroxisome; reconstruction; tomography

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. To date, three proteins have been identified in the fission process in yeast: Fis1, Dnm1, and Mdv1. Fis1 is targeted to the organellar membrane by a C-terminal transmembrane domain (a tail-anchored protein). Dnm1 is a large GTPase thought to act as a mechanoenzyme and belongs to the dynamin superfamily. Mdv1 is an adaptor protein. The current model we are testing is that Fis1 recruits the GTPase Dnm1 to sites of membrane scission and Mdv1 acts to further stimulate Dnm1 assembly, GTP hydrolysis, and membrane scission. We would like to complement our biochemical and NMR/x-ray structural studies with single particle reconstruction and tomography studies on this system. One goal is to build structural models of the fission machinery as a step towards understanding how these proteins couple GTP hydrolysis to membrane scission. We have obtained good expression and purification of Mdv1 constructs fused to maltose binding protein of the N-terminal domain (1-236) and the N-terminal domain with the coiled coil domain (1-302). MBP-Mdv1(1-302) appears to self-assemble, and negative stain EM reveals protofilaments that may arise from MBP, or from Mdv1 itself. This proposal is specifically to image our MBPMdv1(1-302) sample, however, we see a long-term need to work with NCMI on this project as we build up the system in the presence of membranes and other components. We seek to determine the molecular basis for mitochondrial and peroxisome fission, a process essential for organellar function and important in human health.

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。 迄今为止，在酵母的裂变过程中已鉴定出三种蛋白质：Fis1、Dnm1 和 Mdv1。 Fis1 通过 C 端跨膜结构域（尾部锚定蛋白）靶向细胞器膜。 Dnm1 是一种大型 GTP 酶，被认为是一种机械酶，属于动力超家族。 Mdv1 是一种接头蛋白。目前我们正在测试的模型是 Fis1 将 GTPase Dnm1 招募到膜断裂位点，而 Mdv1 则进一步刺激 Dnm1 组装、GTP 水解和膜断裂。我们希望通过对该系统的单粒子重建和断层扫描研究来补充我们的生化和核磁共振/X 射线结构研究。一个目标是建立裂变机制的结构模型，作为了解这些蛋白质如何将 GTP 水解与膜断裂耦合的一步。 我们已经获得了与麦芽糖结合蛋白的 N 端结构域 (1-236) 和带有卷曲螺旋结构域 (1-302) 的 N 端结构域融合的 Mdv1 构建体的良好表达和纯化。 MBP-Mdv1(1-302) 似乎自组装，负染色电镜显示可能由 MBP 或 Mdv1 本身产生的原丝。该提案专门用于对我们的 MBPMdv1(1-302) 样本进行成像，但是，我们认为在该项目上长期需要与 NCMI 合作，因为我们在存在膜和其他组件的情况下构建系统。 我们试图确定线粒体和过氧化物酶体裂变的分子基础，这是细胞器功能所必需的过程，对人类健康也很重要。