• 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Journal of Molecular Biology >Biochemical identification of base and phosphate contacts between Fis and a high-affinity DNA binding site.
【24h】

Biochemical identification of base and phosphate contacts between Fis and a high-affinity DNA binding site.

机译：生化鉴定Fis和高亲和力DNA结合位点之间的碱基和磷酸盐接触。

获取原文
获取原文并翻译 | 示例
           
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

Fis (factor for inversion stimulation) is a nucleoid-associated protein in Escherichia coli and other bacteria that stimulates certain site-specific DNA recombination events, alters DNA topology, and serves as a global gene regulator. DNA binding is central to the functions of Fis and involves a helix-turn-helix DNA binding motif located in the carboxy-terminal region. Specific DNA binding is observed at a number of sites exhibiting poorly related sequences. Such interactions require four critical base pairs positioned -7, -3, +3, and +7 nucleotides relative to the central nucleotide of a 15-bp core-binding site. To further understand how Fis interacts with DNA, we identified the positions of 14 DNA phosphates (based on ethylation interference assays) that are required for Fis binding. These are the 5' phosphates of the nucleotides at positions -8, -7, -6, +1, +2, +3, and +4 relative to the central nucleotide on both DNA strands. Another five phosphates located in the flanking regions from positions +10 through +14 can serve as additional contact sites. Using a combination of biochemical approaches and various mutant Fis proteins, we probed possible interactions between several key Fis residues and DNA bases or phosphates within a high-affinity binding site. We provide evidence in support of interactions between the R85 Fis residue and a highly conserved guanine at position -7 and between T87 and the critical base pairs at -3 and +3. In addition, we present evidence in support of interactions between N84 and the phosphate 5' to the base at +4, between R89 and the -7 phosphate, between T87 and the +3 and +4 phosphates, and between K90 and the +3 phosphate. This work provides functional evidence for some of the most critical interactions between Fis and DNA required for a high binding affinity and demonstrates the large contribution made by numerous phosphates to the stability of the Fis-DNA complex.
机译：Fis（倒转刺激因子）是大肠杆菌和其他细菌中与核苷相关的蛋白，可刺激某些特定位点的DNA重组事件，改变DNA拓扑结构并充当全局基因调节剂。 DNA结合对Fis的功能至关重要，并且涉及位于羧基末端区域的螺旋-转-螺旋DNA结合基序。在表现出不良相关序列的许多位点观察到特异性DNA结合。这样的相互作用需要相对于15bp核心结合位点的中央核苷酸位于-7，-3，+ 3和+7个核苷酸的四个关键碱基对。为了进一步了解Fis如何与DNA相互作用，我们确定了Fis结合所需的14种DNA磷酸盐（基于乙基化干扰分析）的位置。这些是相对于两条DNA链上的中央核苷酸的-8，-7，-6，+ 1，+ 2，+ 3和+4位核苷酸的5'磷酸。位于+10到+14位置的侧翼区域中的另外五种磷酸盐可以用作其他接触位点。使用生化方法和各种突变的Fis蛋白质的组合，我们探测了几个关键的Fis残基与高亲和力结合位点内的DNA碱基或磷酸盐之间的可能相互作用。我们提供证据支持R85 Fis残基与位置-7的高度保守的鸟嘌呤之间以及T87与-3和+3的关键碱基对之间的相互作用。此外，我们提供的证据支持N84与磷酸酯5'到+ 4，R89与-7磷酸酯之间，T87与+3和+4磷酸酯之间，以及K90与+3磷酸酯之间的相互作用。磷酸盐。这项工作为高结合亲和力所需的Fis与DNA之间的一些最关键的相互作用提供了功能证据，并证明了许多磷酸盐对Fis-DNA复合物的稳定性做出了巨大贡献。

著录项

相似文献

  • 外文文献
  • 中文文献
  • 专利
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号