An HXYZ-g HR-Fast-MAS probe for Dramatically Improved Biomolecular Structure Determinations

用于显着改进生物分子结构测定的 HXYZ-g HR-Fast-MAS 探针

• 批准号: 9988618
• 负责人: Francis DAVID Doty
• 金额: $ 64.31万
• 依托单位: DOTY SCIENTIFIC, INC.
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-03-01 至 2020-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9988618
• 关键词: Alzheimer' s Disease; Amyloid beta-Protein; Area; Biological; Biomedical Research; Budgets; COSY; Caliber; Catalysis; Cellular Membrane; Complex; Data; Deposition; Detection; Development; Diffusion; Equipment; Funding; Goals; Health; Inferior; Laboratories; Liquid substance; Magic; Magnetism; Measurement; Membrane Proteins; Metabolism; Methods; Modeling; Molecular; Molecular Weight; Motion; NMR Spectroscopy; Neurology; Nuclear; Nuclear Magnetic Resonance; Performance; Phase; Physiologic pulse; Platelet Factor 4; Price; Proteins; Protocols documentation; Publishing; RF coil; Research Personnel; Resolution; Sampling; Scheme; Side; Solid; Solvents; Sonication; Speed; Structural Protein; Structure; Surface; System; Techniques; Technology; Time; Width; base; data warehouse; design; experimental study; improved; innovation; insight; interest; irradiation; macromolecule; materials science; microwave electromagnetic radiation; next generation; novel; operation; protein structure; solid state nuclear magnetic resonance; structural biology; tool; vibration; virtual

An HXYZ-g HR-Fast-MAS probe for Dramatically Improved Biomolecular Structure Determinations Abstract. Liquid state NMR spectroscopy is arguably one of the best tools for structure determination for soluble proteins. The method provides atomic resolution for modest molecular weight proteins and/or their complexes. The method begins to have difficulty when the molecular weight of the system causes slow molecular motion, which in turn increases the linewidth beyond the point of useful resolution. Solid state NMR (ssNMR) methods have progressed remarkably over the past 15 years to permit improved resolution for these conditions, but they still come well short of the goal of liquid-like resolution on biological macromolecules, such as membrane proteins and the fibrils that are central to Alzheimer’s Disease. The “Holy Grail” in ssNMR would be the ability to successfully utilize the powerful suite of NMR acquisition and automated structure determination protocols developed for solution NMR, which rely on ¹H-detected triple- and quad-resonance ²H-decoupled schemes (as such generally permit 8 or 30 times higher S/N than direct detection, for ¹³C and¹⁵5N respectively) with solid samples of 1-10 mg. The main objective of this Phase II application is to complete the development a four-channel multinuclear ssNMR probe (HXYZ) that has the capability of providing ¹³C/¹⁵N/¹H correlations under ²H decoupling utilizing modest (15 kHz) to fast (> 35 kHz) Magic Angle Spinning (MAS) while detecting ¹H. The resulting resolution, particularly with proposed novel pulse sequences, will be close to that of a typical liquid state experiment on proteins. Four-channel multinuclear probes with gradients have been the workhorse in solution NMR for decades, but they have not been available for ssNMR – they have been perceived to be impractically difficult to design and build. The Phase-I demonstrated feasibility of an H/X/Y/Z narrow-bore (NB) MAS probe based on a novel “single-coil” rf circuit optimized for ¹H detection and suitable for use at fields from 7-31 T. The Phase-II probe will be compatible with automated sample exchange, pulsed-field gradients (PFG), NB magnets, and novel NB microwave irradiation methods for Dynamic Nuclear Polarization (DNP). Calculations suggest ²H J-couplings contribute 5-10 Hz to the remaining ¹H line broadening in rigid proteins, and available data suggest the probe-limited resolution (from thermal gradients and magnetism) in commercially available fast-MAS probes has contributed another 6-25 Hz. A 4-channel MAS probe with order- of-magnitude lower thermal gradients that is capable of 2 Hz ¹H resolution on liquids is expected to enable ¹H linewidths below 0.01 ppm on most of the residues in rigid proteins at 900 MHz and above. The novel circuit will also be tunable to virtually all combinations of interest, such as ¹H/¹³C/²H/¹⁵N, ¹H/³¹P/¹³C/²H, ¹H/³¹P/⁷Li/¹³C, ¹H/²⁷Al/²⁹Si/¹⁷O, and ¹H/¹³C/²⁹Si/¹⁰³Rh, thereby making it also invaluable in such areas as metabolism, neurology, materials science, catalysis, and sustainable energy.

用于显着改进生物分子结构测定的 HXYZ-g HR-Fast-MAS 探针 抽象的。 液态核磁共振波谱可以说是测定可溶性物质结构的最佳工具之一。 蛋白质。该方法为中等分子量的蛋白质和/或其复合物提供原子分辨率。 当系统的分子量导致分子运动缓慢时，该方法开始出现困难， 这反过来又增加了线宽，超出了有用的分辨率点。固态核磁共振 (ssNMR) 方法 在过去 15 年中，在改善这些情况的解决方案方面取得了显着进展，但它们 仍然距离生物大分子（例如膜）类似液体分辨率的目标有很大差距 蛋白质和原纤维是阿尔茨海默病的核心。 ssNMR 中的“圣杯”是能力 成功利用强大的 NMR 采集和自动结构测定协议套件 为溶液 NMR 开发，依赖于 1H 检测的三重和四重共振 2H 解耦方案（如 对于 13C 和 155N），这种方法通常比直接检测的信噪比高 8 或 30 倍 1-10 mg 样品。 该二期应用的主要目标是完成四通道的开发 多核 ssNMR 探针 (HXYZ)，能够在 2H 下提供 13C/15N/1H 相关性 在检测 1H 时利用适度 (15 kHz) 到快速 (> 35 kHz) 魔角旋转 (MAS) 进行去耦。这 由此产生的分辨率，特别是所提出的新型脉冲序列，将接近于典型液体的分辨率 蛋白质国家实验。 具有梯度的四通道多核探针一直是溶液 NMR 的主力 几十年来，但它们还没有可用于 ssNMR——它们被认为是不切实际的困难 设计和建造。第一阶段展示了基于 H/X/Y/Z 窄孔 (NB) MAS 探头的可行性 一种新颖的“单线圈”射频电路，针对 1H 检测进行了优化，适用于 7-31 T 的场。 探头将与自动样品交换、脉冲场梯度 (PFG)、NB 磁铁和 用于动态核极化 (DNP) 的新型 NB 微波辐射方法。 计算表明 ²H J 联轴器对刚性中剩余的 1H 线加宽贡献了 5-10 Hz 蛋白质和可用数据表明探针有限的分辨率（来自热梯度和磁性） 商用快速 MAS 探头又贡献了 6-25 Hz 的频率。一个 4 通道 MAS 探头，具有阶 数量级较低的热梯度能够在液体上实现 2 Hz 1H 分辨率，预计将能够实现 1H 在 900 MHz 及以上频率下，刚性蛋白质中大多数残基的线宽低于 0.01 ppm。新颖的电路 也可调谐到几乎所有感兴趣的组合，例如 1H/13C/2H/15N、1H/31P/13C/2H、1H/31P/7Li/13C、 1H/27Al/2⁹Si/17O 和 1H/13C/29Si/103Rh，从而使其在新陈代谢等领域也具有无价的价值， 神经学、材料科学、催化和可持续能源。