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.

生物分子高场和FAST-MAS SSNMR方法的研究与开发2项目总结 结构与动力(Pi：Hong) 研发2寻求开发新的固态核磁共振技术来确定蛋白质结构 和动力学，并生产用于膜蛋白结构生物学的同位素标记胆固醇。 现代固体核磁共振结构测定方法主要以13C和15N为基础 共振分配和距离测量。然而，13C和13C的低旋磁比 15N将可测量的距离限制在~8？以内，这使得测定蛋白质变得困难 三维褶皱和第四系构造。这项研发的目标1就是克服这一瓶颈 发展基于~(19)F的MAS核磁共振技术测定~(19)F-~(19)F、~(19)F-~(13)C 和19F-1H距离，潜在到2纳米。这些实验将朝着高场、快的方向发展。 MAS条件，以达到结构确定所需的高分辨率和灵敏度。 系统地研究了19F化学位移各向异性对偶极再耦合的影响。 这项研究和开发的目标2是开发13C-2H相关核磁共振技术来测量蛋白质动力学 强磁场，具有比以前更高的角灵敏度。该目标解决了以下限制 基于13C-1H和15N-1H偶极耦合的技术，它们对小电流的敏感度越来越低 振幅运动随着磁场强度和MAS频率的增加而增加。我们将收获 大2H四极耦合探测分子取向的能力 2H自旋和13C或15N自旋之间的极化转移技术，这样我们就可以测量13C 15N分辨~2H四极谱。在这项研发的目标3中，我们将开发生物合成方法 用于高产量生产13C和2H标记的胆固醇，用于膜蛋白结构生物学。通过 我们将与四个推动生物医学项目合作，测试和部署这项研发中的技术 解决膜蛋白和细胞壁领域悬而未决的生物学问题。