Yu C, Bhaskaran R, Chuang L C, Yang C C
Chemistry Department, National Tsing Hua University, Hsinchu, Taiwan, Republic of China.
Biochemistry. 1993 Mar 9;32(9):2131-6. doi: 10.1021/bi00060a002.
The solution conformation of cobrotoxin has been determined by using proton nuclear magnetic resonance spectroscopy. With the combination of various two-dimensional NMR techniques, the 1H-NMR spectrum of cobrotoxin was completely assigned (Yu et al., 1990). A set of 435 approximate interproton distance restraints was derived from nuclear Overhauser enhancement (NOE) measurements. These NOE constraints, in addition to the 29 dihedral angle constraints (from coupling constant measurements) and 26 hydrogen bonding restraints (from the pattern of short-range NOEs), form the basis of 3-D structure determination by the hybrid distance geometry-dynamical simulated annealing method. The 23 structures that were obtained satisfy the experimental restraints, display small deviation from idealized covalent geometry, and possess good nonbonded contacts. Analysis of converged structures indicated that there are two antiparallel beta sheets (double and triple stranded), duly confirming our earlier observations. These are well defined in terms of both atomic root mean square (RMS) differences and backbone torsional angles. The average backbone RMS deviation between the calculated structures and the mean structure, for the beta-sheet regions, is 0.92 A. The mean solution structure was compared with the X-ray crystal structure of erabutoxin b, the homologous protein. This yielded information that both structures resemble each other except at the exposed loop/surface regions, where the solution structure seems to possess more flexibility.
通过使用质子核磁共振光谱法确定了眼镜蛇毒素的溶液构象。结合各种二维核磁共振技术，完全归属了眼镜蛇毒素的1H-NMR谱（Yu等人，1990年）。从核Overhauser增强（NOE）测量中获得了一组435个近似质子间距离限制。这些NOE限制，除了29个二面角限制（来自耦合常数测量）和26个氢键限制（来自短程NOE模式）之外，构成了通过混合距离几何-动力学模拟退火方法确定三维结构的基础。所获得的23个结构满足实验限制，与理想化的共价几何结构偏差较小，并且具有良好的非键接触。对收敛结构的分析表明，存在两个反平行β折叠片层（双链和三链），这充分证实了我们早期的观察结果。这些在原子均方根（RMS）差异和主链扭转角方面都有很好的定义。在β折叠片层区域，计算结构与平均结构之间的平均主链RMS偏差为0.92埃。将平均溶液结构与同源蛋白—— erabutoxin b的X射线晶体结构进行了比较。这得出的信息是，除了在暴露的环/表面区域外，两种结构彼此相似，在该区域溶液结构似乎具有更大的灵活性。
