M. Hong RT&D

洪明 RT

• 批准号: 10224250
• 负责人: Mei Hong
• 金额: $ 5.6万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10224250
• 关键词: 3-Dimensional; Address; Amyloid Fibrils; Anisotropy; Area; Automobile Driving; Binding; Biological; Biosynthetic Proteins; Cell Wall; Chemicals; Cholesterol; Coupling; Disease; Electrons; Frequencies; Harvest; Investigation; Ion Channel; Isotope Labeling; Label; Ligands; Magnetic Resonance; Measurable; Measurement; Measures; Membrane; Membrane Proteins; Methods; Modernization; Molecular; Motion; Nature; Pharmaceutical Chemistry; Physiologic pulse; Production; Protein Dynamics; Proteins; Reagent; Relaxation; Resolution; Site; Structure; Techniques; Technology; Testing; Time; Water; base; beta pleated sheet; experimental study; magnetic field; novel; protein function; protein structure; radio frequency; small molecule; solid state nuclear magnetic resonance; structural biology; three dimensional structure

Project Summary of TR&D 2 – High-field and fast-MAS SSNMR methods for biomolecular structure and dynamics (PI: Hong) TR&D 2 seeks to develop new solid-state NMR technologies to determine protein structure and dynamics and to produce isotopically labeled cholesterol for membrane protein structural biology. Modern solid-state NMR based structure determination approaches focus on 13C and 15N based resonance assignment and distance measurements. However, the low gyromagnetic ratios of 13C and 15N limit the measurable distances to less than ~8 Å, which makes it difficult to determine protein three-dimensional folds and quaternary structures. Aim 1 of this TR&D is to overcome this bottleneck in structure determination by developing 19F-based MAS NMR techniques to measure 19F-19F, 19F-13C and 19F-1H distances, potentially to 2 nm. These experiments will be developed for high field and fast MAS conditions to achieve the high-resolution and sensitivity necessary for structure determination. The impact of 19F chemical shift anisotropy on dipolar recoupling will be investigated systematically. Aim 2 of this TR&D is to develop 13C-2H correlation NMR techniques to measure protein dynamics at high magnetic fields with higher angular sensitivity than before. This aim addresses the limitation of 13C-1H and 15N-1H dipolar-coupling based techniques, which are increasingly less sensitive to small- amplitude motions as the magnetic field strength and MAS frequency increase. We will harvest the power of the large 2H quadrupolar coupling for sensing molecular orientations by developing robust polarization transfer techniques between 2H spins and 13C or 15N spins, so that we can measure 13C and 15N resolved 2H quadrupolar spectra. In aim 3 of this TR&D, we will develop biosynthetic methods for high-yield production of 13C and 2H-labeled cholesterol for membrane protein structural biology. By partnering with four driving biomedical projects, we will test and deploy the technologies in this TR&D to address outstanding biological questions in the areas of membrane proteins and cell walls.

TR&D 2项目总结-生物分子的高场和快速MAS SSNMR方法 结构与动力学（PI：Hong） TR&D 2寻求开发新的固态NMR技术来确定蛋白质结构 和动力学，并产生用于膜蛋白结构生物学的同位素标记胆固醇。 基于现代固态NMR的结构测定方法集中于基于13 C和15 N的结构测定。 谐振分配和距离测量。然而，13 C和14 C的低旋磁比， 15 N限制了可测量的距离小于~8 μ m，这使得难以测定蛋白质 三维褶皱和第四纪构造。本TR&D的目标1是克服这一瓶颈 通过开发基于19 F的MAS NMR技术来测量19 F-19 F、19 F-13 C 和19 F-1H距离，可能达到2nm。这些实验将发展为高场和快速 MAS条件，以实现结构测定所需的高分辨率和灵敏度。 系统地研究了19 F化学位移各向异性对偶极再耦合的影响。 本TR&D的目标2是开发13 C-2 H相关NMR技术，以测量蛋白质动力学， 更高的磁场，具有比以前更高的角灵敏度。这一目标解决了 基于13 C-1H和15 N-1H偶极偶联的技术，其对小分子的灵敏度越来越低。 随着磁场强度和MAS频率的增加，振幅运动。我们将收获 功率的大2 H四极耦合传感分子取向，通过发展鲁棒 在2 H自旋和13 C或15 N自旋之间的极化转移技术，以便我们可以测量13 C 和~（15）N分辨~ 2 H四极谱。在本TR&D的目标3中，我们将开发生物合成方法， 用于膜蛋白结构生物学的13 C和2 H标记胆固醇的高产率生产。通过 我们将与四个主要的生物医学项目合作，测试和部署本TR&D中的技术 以解决膜蛋白和细胞壁领域中悬而未决的生物学问题。