PROTEIN ASSEMBLIES AND METALLOPROTEINS

蛋白质组装体和金属蛋白质

• 批准号: 7960414
• 负责人: Tatyana Polenova
• 金额: $ 20.34万
• 依托单位: UNIVERSITY OF DELAWARE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7960414
• 关键词: Biocompatible Materials; Chloride Peroxidase; Computer Retrieval of Information on Scientific Projects Database; Data; Funding; Grant; Hand; Institution; Investigation; Label; Metalloproteins; Molecular; NMR Spectroscopy; Proteins; Protocols documentation; Research; Research Personnel; Resolution; Resources; Role; Scheme; Solid; Solutions; Source; Structure; Substrate Specificity; United States National Institutes of Health; Vanadium; Vertebral column; X-Ray Crystallography; design; research study; solid state; solid state nuclear magnetic resonance; two-dimensional

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. PROJECT VI: Polenova - Project will investigate structure and conformational dynamics of a 68 kDa- vanadium chloroperoxidase in the solid state using multidimensional NMR spectroscopy. The focus on two major objectives: 1) determine the tertiary structure of chloroperoxidase using solid-state NMR data only; 2) to establish the multiple timescale conformational dynamics in the protein. To accomplish these objectives, three specific aims will be pursued: 1) 13C and 15N resonance assignments of uniformly labeled vanadium chloroperoxidase; 2) determination of long-range tertiary constraints and structure calculation; 3) investigation of multiple timescale conformational dynamics in uniformly and sparsely isotopically enriched protein. These studies would on one hand help elucidate the role of internal conformational dynamics in the substrate specificity of the vanadium haloperoxidases, and on the other, expand the capabilities of solids NMR to structural characterization of large noncrystalline proteins currently intractable by X-ray crystallography or solution NMR. Experimental plan. We completed the specific aim 1 and made significant progress for aim 2. The following experiments were conducted first: 1) heteronuclear 13C-15N two-dimensional correlation experiments to establish backbone resonance assignments, using the double cross polarization (DCP) mixing scheme;1 2) homonuclear two-dimensional 13C-13C correlation experiments to establish the sidechain assignments, using RFDR2 and RAD3 mixing schemes. In the possible case of the insufficient spectral resolution, the three-dimensional NCC protocols were pursued, involving NCACO and NCACB band-selective magnetization transfer schemes. With these experiments in hand, we have to assigned most of the resonances in the protein.

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。 项目VI：POLENOVA - 项目将使用多维NMR光谱法研究固态中68 kDa -vanadium氯氧化酶的结构和构象动力学。 重点是两个主要目标：1）仅使用固态NMR数据确定植物氧化酶的三级结构； 2）在蛋白质中建立多个时间尺度构象动力学。 为了实现这些目标，将追求三个特定的目标：1）1）13C和15N共振分配均匀标记的钒氯氧化酶； 2）确定远程三级约束和结构计算； 3）研究均匀和稀疏的同位素富集蛋白中多个时尺构象动力学。 这些研究一方面将有助于阐明内部构象动力学在钒卤代氧化酶的底物特异性中的作用，另一方面，将固体NMR的能力扩展到当前通过X射线晶体学或溶液NMR进行的大型非晶体蛋白的结构表征。 实验计划。 我们完成了特定的目标1并为目标2取得了重大进展。首先进行了以下实验：1）异核15C-15N二维相关实验，使用双重交叉极化（DCP）混合方案； 1 2）使用SIDECH CORLELAINS建立SIDCECH RAR RAS，使用双交叉极化（DCP）混合方案； 1 2）方案。 在光谱分辨率不足的情况下，采用了三维NCC方案，涉及NCACO和NCACB带相反的磁化转移方案。 借助这些实验，我们必须分配蛋白质中的大多数共振。