Evaluating the binding affinities of NF-κB p50 homodimer to the wild-type and single-nucleotide mutant Ig-κB sites by the unimolecular dsDNA microarray
Evaluating the binding affinities of NF-κB p50 homodimer to the wild-type and single-nucleotide mutant Ig-κB sites by the unimolecular dsDNA microarray
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DOI：
10.1016/s0003-2697(03)00049-6
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发表时间：
2003-05-15
影响因子：
2.9
通讯作者：
Lu, ZH
中科院分区：
文献类型：
--
作者：
Wang, JK;Li, TX;Lu, ZH
本研究采用一种新的方法制备了单分子dsDNA微阵列，研究了NF-κ B p50同源二聚体与野生型和单核苷酸突变型Ig-kappaB位点的结合亲和力。因此评估了Ig-κ B位点的每个核苷酸对于序列特异性p50/Ig-κ B相互作用的重要性。结果表明，不同位置的核苷酸对p50 p50/Ig-kappaB结合相互作用的贡献不同。G（1）、G（2）和C-10对于p50 p50/lg-kappaB结合相互作用是最重要的，并且决定了p50 p50/lg-kappaB相互作用的特异性，用任何其他核苷酸替换可能导致类似的最大结合亲和力损失。相比之下，G（3）、A（4）、T-8和C-9对于p50/lg-kappaB相互作用不太重要，并且调节结合亲和力，其中用变体核苷酸取代可以不同地改变结合亲和力。C-5对p50 p50/lg-kappaB相互作用最不重要，其随机核苷酸交换对p50 p50/lg-kappaB结合亲和力影响很小。在所有可能的单核苷酸突变体中，T-8至C突变可增强p50/lg-kappaB相互作用。T-7的作用与其对称的C-5不同，轴向T-6是高亲和力p50/lg-kappaB相互作用所必需的。单分子dsDNA微阵列为探索DNA结合蛋白与大量DNA靶标的结合亲和力提供了可靠的方法。(C)2003 Elsevier Science（美国）。All rights reserved.
This study investigated the binding affinities of NF-kappaB p50 homodimer to the wild-type and single-nucleotide mutant Ig-kappaB sites by the unimolecular dsDNA microarray which was fabricated with a novel scheme. The importance of each nucleotide of Ig-kappaB site for the sequence-specific p50p50/Ig-kappaB interaction was thus evaluated. The results demonstrate that the nucleotides at different positions contribute differently to the p50p50/Ig-kappaB binding interaction. The G(1), G(2), and C-10 are most important for p50p50/lg-kappaB binding interaction and determine the specificity of p50p50/lg-kappaB interaction, which replacements with any other nucleotide could result in the similarly greatest binding affinity losses. Comparatively, the G(3), A(4), T-8, and C-9 are less important for p50p50/lg-kappaB interaction and regulate the binding affinity, which substitutions with the variant nucleotide could change the binding affinity differently. The C-5 is least important for p50p50/lg-kappaB interaction, the randomized nucleotide exchange of which little affects on p50p50/lg-kappaB binding affinity. Among all possible single-nucleotide mutants, the T-8 to C mutation could strengthen p50p50/lg-kappaB interaction. The T-7 acts differently from its symmetric C-5 and the axial T-6 is necessary for high-affinity p50p50/lg-kappaB interaction. The unimolecular dsDNA microarray provides a reliable method for exploring the binding affinities of DNA-binding proteins with a larger number of DNA targets. (C) 2003 Elsevier Science (USA). All rights reserved.