Kielkopf C L, Ding S, Kuhn P, Rees D C
Division of Biology, California Institute of Technology, Pasadena, CA 99125, USA.
J Mol Biol. 2000 Feb 25;296(3):787-801. doi: 10.1006/jmbi.1999.3478.
The affinity and specificity of a ligand for its DNA site is a function of the conformational changes between the isolated and complexed states. Although the structures of a hydroxypyrrole-imidazole-pyrrole polyamide dimer with 5'-CCAGTACTGG-3' and the trp repressor recognizing the sequence 5'-GTACT-3' are known, the baseline conformation of the DNA site would contribute to our understanding of DNA recognition by these ligands. The 0.74 A resolution structure of a B-DNA double helix, 5'-CCAGTACTGG-3', has been determined by X-ray crystallography. Six of the nine phosphates, two of four bound calcium ions and networks of water molecules hydrating the oligonucleotide have alternate conformations. By contrast, nine of the ten bases have a single, unique conformation with hydrogen atoms visible in most cases. The polyamide molecules alter the geometry of the phosphodiester backbone, and the water molecules mediating contacts in the trp repressor/operator complex are conserved in the unliganded DNA. Furthermore, the multiple conformational states, ions and hydration revealed by this ultrahigh resolution structure of a B-form oligonucleotide are potentially general considerations for understanding DNA-binding affinity and specificity by ligands.
配体与其DNA位点的亲和力和特异性是分离状态与复合状态之间构象变化的函数。虽然已知具有5'-CCAGTACTGG-3'的羟基吡咯-咪唑-吡咯聚酰胺二聚体以及识别序列5'-GTACT-3'的色氨酸阻遏物的结构，但DNA位点的基线构象将有助于我们理解这些配体对DNA的识别。已通过X射线晶体学确定了B-DNA双螺旋5'-CCAGTACTGG-3'的0.74埃分辨率结构。九个磷酸中的六个、四个结合钙离子中的两个以及使寡核苷酸水合的水分子网络具有交替构象。相比之下，十个碱基中的九个具有单一、独特的构象，在大多数情况下氢原子可见。聚酰胺分子改变了磷酸二酯主链的几何形状，并且在色氨酸阻遏物/操纵基因复合物中介导接触的水分子在未结合配体的DNA中得以保留。此外，这种B型寡核苷酸的超高分辨率结构所揭示的多种构象状态、离子和水合作用，可能是理解配体对DNA结合亲和力和特异性的一般考虑因素。
