Stereospecific Effects of Oxygen-to-Sulfur Substitution in DNA Phosphate on Ion Pair Dynamics and Protein-DNA Affinity.
Stereospecific Effects of Oxygen-to-Sulfur Substitution in DNA Phosphate on Ion Pair Dynamics and Protein-DNA Affinity.
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DOI：
10.1002/cbic.201600265
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发表时间：
2016-09-02
期刊：
影响因子：
3.2
通讯作者：
Iwahara, Junji
中科院分区：
文献类型：
--
作者：
Dan Nguyen;Zandarashvili, Levani;White, Mark A.;Iwahara, Junji
DNA磷酸中的氧硫取代通常会增强对DNA结合蛋白的亲和力。我们以前的研究表明，这种效果的硫取代OP 1和OP 2原子是由于熵增益与增强的离子对动力学。在这项工作中，我们研究了单硫取代的OP 1或OP 2原子的DNA磷酸在Lys 57的相互作用位点的双足同源结构域-DNA复合物的立体特异性效应。通过晶体学，我们获得了RP和SP的非对映异构体的硫代磷酸酯和它们与Lys 57的相互作用的结构信息。通过基于荧光的测定，我们发现OP 2原子的硫取代后显着的亲和力增强。通过NMR光谱法，我们发现在OP 2原子的硫取代后，Lys 57侧链NH3+基团的显著移动。这些数据提供了进一步的机制洞察DNA磷酸盐中的氧硫取代的亲和力增强。DNA磷酸中的氧硫取代可以增强蛋白质-DNA结合亲和力。为了获得对这种现象的机理洞察，我们比较了3种不同类型的硫代磷酸酯化对结合亲和力、离子对动力学和结构的影响。
Oxygen-to-sulfur substitutions in DNA phosphate often enhance affinity for DNA-binding proteins. Our previous studies have suggested that this effect of sulfur substitution of both OP1 and OP2 atoms is due to an entropic gain associated with enhanced ion-pair dynamics. In this work, we studied stereospecific effects of single sulfur substitution of either the OP1 or OP2 atom in DNA phosphate at the Lys57 interaction site of the Antennapedia homeodomain-DNA complex. By crystallography, we obtained the structural information on the RP and SP diastereomers of the phosphoromonothioate and their interaction with Lys57. By fluorescence-based assays, we found significant affinity enhancement upon sulfur substitution of the OP2 atom. By NMR spectroscopy, we found significant mobilization of the Lys57 side-chain NH3+ group upon sulfur substitution of the OP2 atom. These data provide further mechanistic insight into the affinity enhancement by oxygen-to-sulfur substitution in DNA phosphate. Oxygen-to-sulfur substitution in DNA phosphate can enhance protein-DNA binding affinity. To gain mechanistic insight into this phenomenon, we compared the impacts of 3 different types of phosphorothioation on binding affinity, ion-pair dynamics, and structure.
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