Nuclear Magnetic Resonance--new Methods And Molecular St

我们扩展了在弱取向条件下通过NMR光谱研究溶液中大分子结构的技术。新的发展集中在一个程序，允许直接纳入小角X射线散射数据到NMR结构测定。虽然计算昂贵，使用一个？一团？将某些肽基团视为固定单点单元的方法，在全原子计算上将该方法加速了几个数量级。应用γ-S晶体蛋白显示出相当好的适合同源X射线结构后，将实验SAXS数据的结构细化。使用SAXS数据被证明是特别有用的分子配合物的研究和研究在溶液条件下的复杂系统的四级结构。 在洗涤剂胶束中的四聚体钾通道KcsA的研究表明，NMR研究相对较大（65 kDa）的膜蛋白本质上是可行的NMR，只要该系统在高温下保持稳定。NMR数据表明，通道/洗涤剂聚集体翻滚为扁球体，反映了通常跨越膜的疏水表面上存在洗涤剂分子。K+结合被认为是不受干扰的洗涤剂的存在下，并没有证据表明，升高，大幅度运动被发现在封闭状态的所谓的选择性过滤器域的通道。 已经开发了新的实验，其允许表征高度不稳定的质子如RNA中的2'羟基质子的位置。这些实验明确地揭示了RNA的A型螺旋区域中的2 'OH质子指向碱基，并且不像文献中先前报道的那样在C3'和碱基区域之间交替。

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.