Spitzfaden C, Braun W, Wider G, Widmer H, Wüthrich K
Institut für Molekularbiologie und Biophysik, Eidgenössische Technische Hochschule-Hönggerberg, Zürich, Switzerland.
J Biomol NMR. 1994 Jul;4(4):463-82. doi: 10.1007/BF00156614.
The three-dimensional NMR solution structure of the cyclophilin A (Cyp)-cyclosporin A (CsA) complex was determined, and here we provide a detailed description of the analysis of the NMR data and the structure calculation. Using 15N- and 13C-resolved three- and four-dimensional [1H,1H]-nuclear Overhauser enhancement (NOE) spectroscopy with uniformly isotope-labeled Cyp in the complex, a final data set of 1810 intra-Cyp, 107 intra-CsA and 63 intermolecular NOE upper distance constraints was collected as input for the structure calculation with the program DIANA. A group of DIANA conformers, selected by a previously described analysis of the dependence of the maximal root-mean-square deviation (rmsd) among the individual conformers on the residual target function value, was subjected to energy refinement with the program FANTOM. The 22 best energy-refined conformers were then used to represent the solution structure. The average rmsd relative to the mean structure of these 22 conformers is 1.1 A for the backbone atoms of all residues of the complex. The molecular architecture of Cyp in the Cyp-CsA complex includes an eight-stranded antiparallel beta-barrel, which is closed on each side by an amphipathic helix. CsA is bound in a cavity formed by part of the barrel surface and four loops with nonregular secondary structure. Comparison of this structure with structures of Cyp-CsA and other Cyp-peptide complexes determined by different approaches shows extensive similarities.
确定了亲环蛋白A（Cyp）-环孢菌素A（CsA）复合物的三维核磁共振溶液结构，在此我们详细描述了核磁共振数据的分析和结构计算过程。在复合物中使用均匀同位素标记的Cyp进行15N和13C分辨的三维和四维[1H,1H]-核Overhauser增强（NOE）光谱分析，收集了1810个Cyp内部、107个CsA内部和63个分子间NOE上距离约束的最终数据集，作为使用DIANA程序进行结构计算的输入。通过先前描述的对各个构象体之间最大均方根偏差（rmsd）对残余目标函数值的依赖性分析选择的一组DIANA构象体，使用FANTOM程序进行能量优化。然后使用22个最佳能量优化构象体来代表溶液结构。相对于这22个构象体的平均结构，复合物所有残基的主链原子的平均rmsd为1.1埃。Cyp-CsA复合物中Cyp的分子结构包括一个八链反平行β桶，其两侧由两亲性螺旋封闭。CsA结合在由桶表面的一部分和四个具有非规则二级结构的环形成的腔内。将该结构与通过不同方法确定的Cyp-CsA和其他Cyp-肽复合物的结构进行比较，发现有广泛的相似性。
