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

9316161 Schaefer博士建议进行新的固态核磁共振实验，以确定蛋白质与大分子不可结晶的复合体的结构，包括昆虫和植物细胞壁中的蛋白质-蛋白质交联体、蛋白质-低聚糖复合体以及蛋白质-脱氧核糖核酸复合体。他在过去五年中为准确确定核间距离(以及几何和结构)而开发的通用旋转回波双共振(REDOR)和转移回波双共振(TEDOR)分析方法，以及他建议开发的新的组合TEDOR-REDOR和TEDOR-TEDOR方法，都是直接适用于大型络合物分析的技术。固体核磁共振对这类工作的敏感性已经在大蛋白质的实际应用中得到了证实。Schaefer博士现在例行地对嵌入在离子玻璃中的50-kD标记蛋白质复合体的1微克分子进行远距离测量，这种蛋白质量通常用于传统的溶液状态1H多维核磁共振。红外线固体核磁共振的实际应用可以带来新的科学和新的生物技术，后者包括农药、除草剂和药物的生物合成设计。Schaefer博士计划通过核磁共振检查昆虫和植物细胞壁中的蛋白质-蛋白质交联链、蛋白质-低聚碳氢化合物复合体和蛋白质-DNA复合体。这些络合物不能用其他技术来分析，因为它们太大了，或者是太不均匀的基质的一部分。他还建议使用原子力显微镜(AFM)来表征一些我们通过核磁共振检查的相同蛋白质复合体的全球结构。锚定在玻璃状聚合物表面上的蛋白质复合体将用非微扰的“敲击模式”原子力显微镜进行检查。对相同蛋白质系统的核磁共振和原子力显微镜实验的结合将提供内部校准(自我一致性检查)，并应导致新的结构洞察力。***

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. ***