800 MHz Solid-State NMR Spectrometer for Biomacromolecular Structure and Dynamics

用于生物大分子结构和动力学的 800 MHz 固态核磁共振波谱仪

• 批准号: 8334722
• 负责人: Christopher P Jaroniec
• 金额: $ 200万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8334722
• 关键词: Amyloid Fibrils; Biological; Biological Phenomena; Cell Nucleus; Complex; DNA Repair; Data; Diagnostic radiologic examination; Disease; Elements; Equipment; Frequencies; Funding; Gene Expression Regulation; Health; Human; Label; Magic; Measurement; Methodology; Methods; Molecular Weight; Nucleic Acids; Nucleosomes; Ohio; Pathogenesis; Peptides; Play; Preparation; Prions; Proteins; Protocols documentation; Relative (related person); Research Personnel; Research Project Grants; Resolution; Role; Sampling; Signal Transduction; Site; Solutions; Structure; System; Training; United States National Institutes of Health; Universities; X-Ray Crystallography; cold temperature; instrument; macromolecule; magnetic field; molecular mass; novel; operation; protein structure; research study; restraint; solid state nuclear magnetic resonance; success

DESCRIPTION (provided by applicant): We propose to acquire a state-of-the-art 800 MHz wide-bore solid-state NMR spectrometer for studies of complex biomacromolecular assemblies including prion protein amyloid fibrils and large nucleosome arrays. These systems play a major role in diverse biological phenomena such as the pathogenesis of protein conformational diseases, gene regulation and DNA repair, and are of fundamental importance to human health. Most samples employed in these studies exist natively as high-molecular weight non-crystalline or fibrous aggregates. Since such preparations are insoluble and lack long-range order they are not readily interrogated by conventional X-ray crystallography or solution-state NMR methods. On the other hand, they are ideal targets for analysis by solid-state NMR. The proposed instrument will be equipped for advanced multidimensional magic-angle spinning NMR experiments, that will enable the resolution and assignment of 1H, 13C and 15N signals throughout 13C,15N-labeled proteins within complexes having molecular masses up to ~400 kDa. The instrument will also permit determination of long-range interatomic 1H-1H, 13C-13C and 13C-15N distance restraints up to ~5-8 ¿, as well as 15N-Cu2+ and 13C-Cu2+ distances up to ~20-25 ¿ in protein molecules modified at specific sites with paramagnetic Cu2+-chelating tags. Distance restraints of this type are a key ingredient of solid-state NMR structure determination protocols. Moreover, the instrument will provide the capability to readily detect 31P, 2H and 17O nuclei in peptides, proteins and nucleic acids for structural and dynamic measurements, with simple adjustments of the standard probe tuning elements. The novel capabilities of the proposed instrument, which are critical to the success of the proposed experiments, include significantly enhanced sensitivity and resolution relative to low- to moderate-field spectrometers, excellent long-term operational stability for 2D, 3D and 4D NMR experiments, extended operation at low temperatures, and rapid magic-angle spinning at frequencies ¿ 40 kHz. The proposed instrument will support ongoing projects among a core group of investigators at The Ohio State University and Case Western Reserve University in Cleveland, OH with NIH R01 funding, and also enable the application of ultrahigh-field magic-angle-spinning solid-state NMR to research projects at Ohio State which can significantly benefit from this methodology but have not exploited it as of yet due to lack of equipment. The PI has extensive training in advanced magic-angle-spinning solid-state NMR applied to biological macromolecules and has developed multidimensional NMR methods for protein assignments, three-dimensional protein structure refinement and rapid acquisition of protein NMR data with enhanced sensitivity. These and other new methodologies will be implemented on the new 800 MHz NMR instrument to enable studies of biomacromolecular systems that are difficult or impossible to investigate at lower magnetic fields.

描述（由申请人提供）：我们建议购买一台最先进的 800 MHz 宽口径固态核磁共振波谱仪，用于研究复杂的生物大分子组装体，包括朊病毒蛋白淀粉样原纤维和大型核小体阵列。这些系统发挥着重要作用。它们在蛋白质构象疾病的发病机制、基因调控和 DNA 修复等多种生物现象中发挥着重要作用，并且对这些研究中使用的大多数样本具有根本重要性。另一方面，由于此类制剂不溶且缺乏长程有序，因此它们本身以高分子量非晶态或纤维状聚集体形式存在，因此不易通过常规 X 射线晶体学或溶液态 NMR 方法进行研究。拟议的仪器将配备先进的多维魔角旋转核磁共振实验，从而实现 1H、13C 和 15N 信号的解析和分配。该仪器还可以测定分子量高达约 400 kDa 的复合物中的 13C、15N 标记蛋白质，还可以测定高达约 5-8 ¿ 的长程原子间 1H-1H、13C-13C 和 13C-15N 距离限制。 ，以及 15N-Cu2+ 和 13C-Cu2+ 距离可达 ~20-25 ¿这种类型的距离限制是固态 NMR 结构测定方案的关键组成部分，此外，该仪器还能够轻松检测 31P、2H 和 17O 核。通过简单调整标准探针调谐元件，可以对肽、蛋白质和核酸进行结构和动态测量，该仪器的新颖功能对于所提出的实验的成功至关重要，包括显着增强。相对于中低场光谱仪的灵敏度和分辨率，2D、3D 和 4D NMR 实验的出色长期操作稳定性，低温下的延长操作以及频率下的快速魔角旋转 ¿ 40 kHz 的仪器将支持俄亥俄州立大学和俄亥俄州克利夫兰凯斯西储大学的核心研究小组正在进行的项目，并获得 NIH R01 的资助，并支持超高场魔角旋转固体的应用。俄亥俄州立大学的国家核磁共振研究项目可以从这种方法中受益匪浅，但由于缺乏设备，尚未对其进行利用。 PI 在应用于生物大分子的先进魔角旋转固态核磁共振方面接受了广泛的培训。和开发了多维 NMR 方法，用于蛋白质分配、三维蛋白质结构细化和快速采集具有增强灵敏度的蛋白质 NMR 数据。这些和其他新方法将在新型 800 MHz NMR 仪器上实施，以实现困难的生物大分子系统的研究。或者不可能在较低的磁场下进行研究。

项目成果

Rapid Quantitative Measurements of Paramagnetic Relaxation Enhancements in Cu(II)-Tagged Proteins by Proton-Detected Solid-State NMR Spectroscopy.

通过质子检测固态核磁共振光谱快速定量测量 Cu(II) 标记蛋白质的顺磁弛豫增强。

• 发表时间: 2017-12-07
• 作者: Mukhopadhyay, Dwaipayan;Nadaud, Philippe S;Shannon, Matthew D;Jaroniec, Christopher P
• 通讯作者: Jaroniec, Christopher P

Histone H4 Tails in Nucleosomes: a Fuzzy Interaction with DNA.

核小体中的组蛋白 H4 尾部：与 DNA 的模糊相互作用。

• 发表时间: 2021
• 作者: Rabdano, Sevastyan O;Shannon, Matthew D;Izmailov, Sergei A;Gonzalez Salguero, Nicole;Zandian, Mohamad;Purusottam, Rudra N;Poirier, Michael G;Skrynnikov, Nikolai R;Jaroniec, Christopher P
• 通讯作者: Jaroniec, Christopher P