Jiang Jiansheng, Sheng Jia, Carrasco Nicolas, Huang Zhen
Department of Chemistry, Georgia State University, Atlanta, GA 30302, USA.
Nucleic Acids Res. 2007;35(2):477-85. doi: 10.1093/nar/gkl1070. Epub 2006 Dec 14.
The high-resolution structure of the DNA (5'-GTGTACA-C-3') with the selenium derivatization at the 2'-position of T2 was determined via MAD and SAD phasing. The selenium-derivatized structure (1.28 A resolution) with the 2'-Se modification in the minor groove is isomorphorous to the native structure (2.0 A). To directly compare with the conventional bromine derivatization, we incorporated bromine into the 5-postion of T4, determined the bromine-derivatized DNA structure at 1.5 A resolution, and found that the local backbone torsion angles and solvent hydration patterns were altered in the structure with the Br incorporation in the major groove. Furthermore, while the native and Br-derivatized DNAs needed over a week to form reasonable-size crystals, we observed that the Se-derivatized DNAs grew crystals overnight with high-diffraction quality, suggesting that the Se derivatization facilitated the crystal formation. In addition, the Se-derivatized DNA sequences crystallized under a broader range of buffer conditions, and generally had a faster crystal growth rate. Our experimental results indicate that the selenium derivatization of DNAs may facilitate the determination of nucleic acid X-ray crystal structures in phasing and high-quality crystal growth. In addition, our results suggest that the Se derivatization can be an alternative to the conventional Br derivatization.
通过多波长反常散射（MAD）和单波长反常散射（SAD）相位法确定了在T2的2'-位进行硒衍生化的DNA（5'-GTGTACA-C-3'）的高分辨率结构。在小沟中具有2'-Se修饰的硒衍生化结构（分辨率为1.28 Å）与天然结构（2.0 Å）是同晶型的。为了与传统的溴衍生化直接比较，我们将溴掺入T4的5-位，以1.5 Å分辨率确定了溴衍生化的DNA结构，发现随着Br掺入大沟，该结构中的局部主链扭转角和溶剂水合模式发生了改变。此外，虽然天然DNA和Br衍生化的DNA需要一周多的时间才能形成尺寸合适的晶体，但我们观察到Se衍生化的DNA在一夜之间就长出了具有高衍射质量的晶体，这表明Se衍生化促进了晶体形成。此外，Se衍生化的DNA序列在更广泛的缓冲条件下结晶，并且通常具有更快的晶体生长速率。我们的实验结果表明，DNA的硒衍生化可能有助于在相位分析和高质量晶体生长中确定核酸的X射线晶体结构。此外，我们的结果表明，Se衍生化可以替代传统的Br衍生化。
