NUCLEAR MAGNETIC RESONANCE--NEW METHODS AND MOLECULAR STRUCTURE DETERMINATION

核磁共振方法在结构测定中的适用性 大于15kD的蛋白质正在研究中。方法 与那些应用于小蛋白的不同之处在于 减少与增加的数量相关的频谱拥挤 分子中的氢。事实证明， 蛋白质中所有不可交换质子的过氚化合 葡萄球菌核酸酶(18kD)可显著改善 允许的可交换酰胺质子的光谱质量 三个α螺旋区的识别。毫不含糊 然后对酰胺质子的位置进行了特定的共振指定 通过使用新的13C1-15N双标记方法获得。 提出了一种测量1H-1HJ的有效方法 大分子中的偶联，而这些偶联通常不是 可以直接观察到。该方法已应用于DNA 十二聚体d(CGCGAATTCGCG)2.测量联轴器的分析 表明所有脱氧核糖都必须经历快速 在至少两个不同的 几何、N型和S型构象。时间的一部分 每种构象中存在的任何脱氧核糖都随其 在十二聚体中的位置，平均而言，含有嘧啶 N-特征(10-20%)比嘌呤(0-20%)多的核苷酸 10%)。

The applicability of NMR methods for structure determination of proteins larger than 15 kD is being investigated. Approaches different from those applied to small proteins are needed to decrease spectral crowding associated with the increased number of hydrogens in the molecule. lt has been demonstrated that perdeuteration of all non-exchangeable protons in the protein Staphylococcal Nuclease (18 kD) results in a large improvement of spectral quality of the exchangeable amide protons, permitting identification of the three alpha helical regions. Unambiguous site specific resonance assignments of the amide protons were then obtained by using a novel 13C1-15N double labeling approach. An efficient method has been developed for measuring 1H-1H J couplings in macromolecules where these couplings normally are not directly observable. The method has been applied to the DNA dodecamer d(CGCGAATTCGCG)2. Analysis of the measured couplings indicates that all deoxyribose sugars must undergo rapid conformational transitions between a minimum of two different geometries, a N-type and an S-type conformer. The fraction of time any deoxyribose exists in each conformation varies with its position in the dodecamer with, on average, the pyrimidine nucleotides having more N-character (10-20%) than the purines (0- 10%).