Nuclear Magnetic Resonance--new Methods And Molecular St

核磁共振--新方法与分子研究

• 批准号: 7336246
• 负责人: Ad - Bax
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7336246
• 关键词: Nuclear; Magnetic; Resonance; new; Methods

We have extended the technology for studying macromolecular structure in solution by NMR spectroscopy under weakly aligning conditions. New developments focus on a procedure that permits direct incorporation of small angle X-ray scattering data into NMR structure determination. Although computationally expensive, use of a ?glob? approach, which treats certain peptide groups as fixed single point units, accelerates the method by several orders of magnitude over a full atom calculation. Application to gamma-S crystallin showed a considerably better fit to homologous X-ray structure upon incorporation of experimental SAXS data in the structure refinement. Use of SAXS data is proving particularly useful for the study of molecular complexes and for studying quaternary structure of complex systems under solution conditions. Study of the tetrameric potassium channel KcsA in detergent micelles indicates that NMR studies of relatively large (65 kDa) membrane proteins intrinsically is feasible by NMR, provided that the system remains stable at elevated temperature. NMR data indicate that the channel/detergent aggregate tumbles as an oblate spheroid, reflecting the presence of detergent molecules on the hydrophobic surface that normally spans the membrane. K+ binding is found not to be disturbed by the presence of detergent, and no evidence for elevated, large amplitude motions is found in the closed state of the so-called selectivity filter domain of the channel. Novel experiments have been developed that permit characterization of the position of highly labile protons such as 2' hydoxyl protons in RNA. These experiments reveal unambiguously that the 2'OH proton in A-form helical regions of RNA points towards the base, and does not alternate between the C3' and base regions, as reported earlier in the literature.

我们已经扩展了在弱排列条件下通过NMR光谱研究在溶液中研究大分子结构的技术。新的发展集中在一个程序上，该程序允许将小角度X射线散射数据直接融合到NMR结构确定中。虽然计算昂贵，但使用“地球仪？将某些肽组视为固定单点单位的方法，在完整的原子计算中通过几个数量级加速了该方法。在将实验性SAXS数据掺入结构细化后，应用于γ-S Crystallin在同源X射线结构上表现出相当大的拟合。事实证明，SAXS数据的使用对于研究分子复合物的研究以及在溶液条件下研究复合系统的第四纪结构特别有用。 对洗涤剂胶束中四聚体钾通道KCSA的研究表明，NMR本质上对相对大（65 kDa）膜蛋白的NMR研究本质上是可行的，前提是该系统在升高温度下保持稳定。 NMR数据表明，通道/洗涤剂聚集体掉落为块状球体，反映了通常跨膜的疏水表面上的洗涤剂分子的存在。发现K+结合不会受到洗涤剂的存在的干扰，并且在所谓的选择性滤波器的封闭状态中没有发现升高，较大的振幅运动的证据。 已经开发了新的实验，可以表征高度不稳定质子的位置，例如RNA中的2'羟基质子。这些实验明确地表明，RNA的A形螺旋区域中的2'OH质子指向碱基，并且在文献早期报道的那样，在C3'和基础区域之间不交替。