Yamakage S I, Maltseva T V, Nilson F P, Földesi A, Chattopadhyaya J
Department of Bioorganic Chemistry, University of Uppsala, Sweden.
Nucleic Acids Res. 1993 Nov 11;21(22):5005-11. doi: 10.1093/nar/21.22.5005.
The concept of the 1H-NMR window has been developed and examined through a comparative study of NOESY spectra of a self-complementary Dickerson's dodecamer (I) [5'd(5C6G7C8G9A10A11T12T13C-14G15C16G)2(3')], a self-complementary 20-mer (II) [(5'd(1C2G3C4G5C6G7C8G9A10A11T12T13C14G15C16G17C18G19C20G)2(3')] in which the core part consists of the same Dickerson's dodecamer sequence with the flanking CGCG residues at both 3' and 5'-ends, and the partly-deuteriated (shown by underlined CGCG residues at both 3' and 5'-ends) analogous duplex (III) [5'd(1C2G3C4G5C6G7C8G9A10A11T12T13C14G15C16G17C18G19C20G)2(3')] in which the core 5C to 16G part (i.e. 1H-NMR window) consists of the natural Dickerson's dodecamer sequence. A comparison of their NOESY spectra clearly demonstrates that the severe overlap of proton resonances in the larger DNA duplex (II) has been successfully reduced in the partly-deuterated duplex (III) as a result of specific incorporations of the sugar-deuteriated nucleotide residues in the latter [stereospecific > 97 atom % 2H enrichment at H2', H2'' and H3' sites, approximately 85 atom % 2H enrichment at H4' and approximately 20 atom % 2H enrichment at H1' (see refs. 10 and 11) in the 20-mer duplex (III)]. These simplifications of the resonance overlap by the deuteriation approach have enabled unequivocal chemical shift assignments and extraction of the quantitative NOE data in the 1H-NMR window part of duplex (III). A comparison of the 12-nucleotide long 1H-NMR window in (III) with that of the 12-mer duplex (I) also shows the scope of studying the changes in conformation and dynamics of the core 12-mer region in (III) which result from the increase of molecular weight due to the DNA chain extension. It is noteworthy that such a study is clearly impossible for the natural 20-mer (II) because of the inherent problem of the overlap of resonances.
通过对自互补的迪克森十二聚体（I）[5'd(5C6G7C8G9A10A11T12T13C - 14G15C16G)2(3')]、自互补的20聚体（II）[(5'd(1C2G3C4G5C6G7C8G9A10A11T12T13C14G15C16G17C18G19C20G)2(3')]（其核心部分由相同的迪克森十二聚体序列组成，在3'和5'端带有侧翼CGCG残基）以及部分氘代的（3'和5'端带下划线的CGCG残基表示）类似双链体（III）[5'd(1C2G3C4G5C6G7C8G9A10A11T12T13C14G15C16G17C18G19C20G)2(3')]（其核心的5C至16G部分，即1H - NMR窗口，由天然的迪克森十二聚体序列组成）的NOESY谱进行比较研究，1H - NMR窗口的概念得以发展和检验。对它们的NOESY谱进行比较清楚地表明，由于在后者中糖 - 氘代核苷酸残基的特异性掺入（在20聚体双链体（III）的H2'、H2''和H3'位点立体特异性>97原子%的2H富集，在H4'位点约85原子%的2H富集，在H1'位点约20原子%的2H富集（见参考文献10和11）），较大DNA双链体（II）中质子共振的严重重叠在部分氘代双链体（III）中已成功减少。通过氘代方法对共振重叠的这些简化使得在双链体（III）的1H - NMR窗口部分能够明确进行化学位移归属并提取定量的NOE数据。将（III）中12个核苷酸长的1H - NMR窗口与12聚体双链体（I）的进行比较，也显示了研究（III）中核心12聚体区域由于DNA链延伸导致分子量增加而引起的构象和动力学变化的范围。值得注意的是，由于共振重叠这一固有问题，对于天然的20聚体（II）显然无法进行这样的研究。
