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首页> 外文期刊>Journal of Molecular Biology >Distortion of DNA junctions imposed by the binding of resolving enzymes: A fluorescence study
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Distortion of DNA junctions imposed by the binding of resolving enzymes: A fluorescence study

机译：解析酶结合引起的DNA连接扭曲：一项荧光研究

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摘要

Junction-resolving enzymes are nucleases that are specific for the structure of the four-way DNA junction. The binding of RuvC of Escherichia coli and Hjc of Sulfolobus solfataricus can be followed by an increase in the fluorescence anisotropy of Cy3 terminally attached to one of the helical arms of a four-way junction. By contrast, there was no change in fluorescein anisotropy with the binding of single dimers of these proteins. Fluorescence resonance energy transfer has therefore been used between fluorescein and Cy3 fluorophores attached to the ends of helical arms to analyse the global structure of the junction on protein binding. The results indicate that both enzymes induce a marked change in the global DNA conformation on the binding of a single dimer. The structure of the protein-junction complexes is independent of the presence or absence of divalent metal ions, unlike that of the protein-free junction. The structures of the RuvC and Hjc complexes are different, but both represent a significant opening of the structure compared to the stacked X-structure of the protein-free junction in the presence of magnesium ions. This protein-induced opening is likely to be important in the function of these enzymes.
机译：连接解析酶是对四向DNA连接结构具有特异性的核酸酶。大肠杆菌的RuvC与Sulfolobus solfataricus的Hjc结合后，可能会增加末端连接至四向连接的一个螺旋臂的Cy3的荧光各向异性。相比之下，荧光素各向异性与这些蛋白质的单个二聚体的结合没有变化。因此，已在连接至螺旋臂末端的荧光素和Cy3荧光团之间使用了荧光共振能量转移，以分析蛋白质结合处连接的整体结构。结果表明，两种酶在单个二聚体的结合上诱导总体DNA构象的显着变化。与无蛋白连接不同，蛋白连接复合物的结构独立于二价金属离子的存在与否。 RuvC和Hjc配合物的结构不同，但与存在镁离子的无蛋白连接的堆叠X结构相比，两者都表示该结构的显着开口。蛋白质诱导的开放可能在这些酶的功能中很重要。

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