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首页> 外文期刊>Journal of Molecular Biology >FOOTPRINT ANALYSIS OF M.SSSL AND M.HHAL METHYLTRANSFERASES REVEALS EXTENSIVE INTERACTIONS WITH THE SUBSTRATE DNA BACKBONE
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FOOTPRINT ANALYSIS OF M.SSSL AND M.HHAL METHYLTRANSFERASES REVEALS EXTENSIVE INTERACTIONS WITH THE SUBSTRATE DNA BACKBONE

机译：M.SSSL和M.Halal甲基转移酶的足印分析显示与基质DNA骨架的广泛相互作用

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摘要

The interactions of the CpG methyltransferases M.Sssl and M.Hhal (GCGC) with substrate DNA were investigated using three different footprinting techniques. The two structurally related enzymes displayed similar specific and non-specific contacts with DNA while bound to their target sequences. DNase I footprinting implicated a region of 18 to 21 base-pairs with which these enzymes interact. Dimethylsulfate protection experiments mostly revealed specific base interactions; each enzyme was shown to interact predominantly with bases at its recognition site in the major groove. However, hydroxyl radical footprints demonstrated extensive interactions with the sugar-phosphate backbone on both strands of the DNA substrate. Both enzymes protected a 16 nucleotide region, in a staggered fashion, covering 9 to 10 nucleotides on each strand. The protected regions, extending for almost a full turn of DNA on each strand, were offset by 6 to 7 nucleotides in the 5' direction, placing both regions on the same face of the double helix, bracketing the major groove. The results suggest that these methyltransferases straddle the major groove from the backbone, but protrude into the groove only to specifically interact with their recognition sites. The sequence-independent interactions observed on the sugar-phosphate backbone may explain the ability of the enzymes to recognize a small sequence, as well as their processive mode of action. [References: 17]
机译：使用三种不同的足迹技术研究了CpG甲基转移酶M.Sssl和M.Hhal（GCGC）与底物DNA的相互作用。两种结构相关的酶在与它们的靶序列结合时表现出与DNA相似的特异性和非特异性接触。 DNase I足迹涉及这些酶相互作用的18至21个碱基对的区域。硫酸二甲酯的保护实验大多揭示了特定的碱基相互作用;每种酶显示出主要与主要沟中识别位点的碱基相互作用。然而，羟基自由基足迹表明与DNA底物的两条链上的糖-磷酸骨架之间存在广泛的相互作用。两种酶均以交错方式保护16个核苷酸区域，每条链覆盖9至10个核苷酸。在每条链上延伸几乎一整段DNA的受保护区域在5'方向上被6到7个核苷酸偏移，将两个区域都放在双螺旋的同一面上，并在大槽之间加了括号。结果表明，这些甲基转移酶跨越主链的主要沟，但仅突入沟中以与其识别位点特异性相互作用。在糖-磷酸骨架上观察到的与序列无关的相互作用可能解释了酶识别小序列的能力，以及它们的过程性作用方式。 [参考：17]

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