Shapes of Protein-Macromolecule Complexes by Solid-State Nuclear Magnetic Resonance

通过固态核磁共振研究蛋白质-大分子复合物的形状

• 批准号: 9316161
• 负责人: Jacob Schaefer
• 金额: $ 36万
• 依托单位: Washington University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-04-01 至 1997-09-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9316161&HistoricalAwards=false
• 关键词: Shapes; Protein; Macromolecule; Complexes; Solid

9316161 Schaefer Dr. Schaefer proposes new solid-state NMR experiments to determine the structures of big, non-crystallizeable complexes of proteins with macromolecules, including protein-protein cross-links in insect and plant cell walls, protein-oligosaccahride complexes, and protein-DNA complexes. The versatile rotational-echo double- resonance (REDOR) and transferred-echo double-resonance (TEDOR) analytical methods that he has developed in the last five years for the accurate determination of internuclear distances (and hence geometry and structure), and the new combined TEDOR-REDOR and TEDOR-TEDOR methods which he proposes to develop, are techniques that are directly applicable to the analysis of big complexes. The sensitivity of solid-state NMR for this kind of work already has been established in practical applications on big proteins. Dr. Schaefer now routinely performs long-range distance measurements on one umole of a 50-kD labeled protein complex embedded in an ionic glass, an amount of protein commonly used for conventional solution-state 1H multi-dimensional NMR. %%% The practical applications of REDOR solid-state NMR could lead both to new science and new biotechnology, the latter including the biorational design of pesticides, herbicides, and drugs. Dr. Schaefer plans to examine by NMR protein-protein cross-links in insect and plant cell walls, protein-oligosacahride complexes, and protein-DNA complexes. These complexes cannot be analyzed by other techniques because they are too big or are part of too heterogeneous a matrix. He also proposes to use atomic force microscopy (AFM) to characterize the global structure of some of the same protein complexes that we examine by NMR. Protein complexes anchored on the undulations of glassy polymer surfaces will be examined by non-perturbative "tapping-mode" AFM. The combination of NMR and AFM experiments on the same protein systems will provide an internal calibration (a self consistency check) and should lead to new structural insights. ***

9316161 Schaefer Schaefer博士提出了新的固态核磁共振实验，以确定蛋白质与大分子的大型不可结晶复合物的结构，包括昆虫和植物细胞壁中的蛋白质-蛋白质交联、蛋白质-低聚糖复合物和蛋白质-DNA复合物。 在过去五年中，他开发了多功能的旋转回波双共振（REDOR）和转移回波双共振（TEDOR）分析方法，用于精确测定核间距离（以及几何形状和结构），以及他提议开发的新的TEDOR-REDOR和TEDOR-TEDOR组合方法，这些技术直接适用于分析大型络合物。 固体核磁共振对这类工作的灵敏度已经在大蛋白质的实际应用中得到了证实。 Schaefer博士现在常规地对嵌入离子玻璃中的50 kD标记蛋白质复合物的1 umole进行远程距离测量，这是常规溶液状态1H多维NMR常用的蛋白质量。 REDOR固体核磁共振的实际应用可能会导致新的科学和新的生物技术，后者包括杀虫剂，除草剂和药物的生物合理设计。 Schaefer博士计划通过核磁共振研究昆虫和植物细胞壁中的蛋白质-蛋白质交联，蛋白质-寡糖复合物和蛋白质-DNA复合物。 这些复合物不能通过其他技术进行分析，因为它们太大或者是太异质的基质的一部分。 他还建议使用原子力显微镜（AFM）来表征我们通过NMR检查的一些相同蛋白质复合物的全局结构。 锚定在玻璃状聚合物表面波动上的蛋白质复合物将通过非微扰“敲击模式”原子力显微镜进行检查。 在相同蛋白质系统上的NMR和AFM实验的组合将提供内部校准（自我一致性检查）， 应该会带来新的结构性见解。 ***