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15 kD的蛋白质正在研究中。方法 与应用于小蛋白质的不同， 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全氘化 葡萄球菌核酸酶（18 kD）导致大幅度改善 可交换酰胺质子的光谱质量，允许 三个α螺旋区域的鉴定。 明确 酰胺质子的位点特异性共振归属， 通过使用新的13 C1 - 15 N双标记方法获得。 本文提出了一种测量~ 1H-~ 1H J的有效方法 在大分子中的偶联，这些偶联通常不 直接观察。 该方法已应用于DNA 十二聚体d（CGCGAATTCGCG）2. 测量耦合的分析 表明所有的脱氧核糖糖必须经历快速的 最少两种不同的构象之间的构象转变 几何构型、N型和S型构象异构体。 的时间分数 任何脱氧核糖存在于每个构象不同， 在十二聚体中的位置，平均而言， 具有比嘌呤（0-20%）多的N-特征的核苷酸（10-20%）， 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%).