DNA-binding proteins play an important role in gene regulation and cellular function. The transcription factors MarA and Rob are two homologous members of the AraC/XylS family that regulate multidrug resistance. They share a common DNA-binding domain, and Rob possesses an additional C-terminal domain that permits binding of low-molecular weight effectors. Both proteins possess two helix-turn-helix (HTH) motifs capable of binding DNA; however, while MarA interacts with its promoter through both HTH-motifs, prior studies indicate that Rob binding to DNA via a single HTH-motif is sufficient for tight binding. In the present work, we perform microsecond time scale all-atom simulations of the binding of both transcription factors to different DNA sequences to understand the determinants of DNA recognition and binding. Our simulations characterize sequence-specific changes in dynamical behavior upon DNA binding, showcasing the role of Arg40 of the N-terminal HTH-motif in allowing for specific tight binding. Finally, our simulations demonstrate that an acidic C-terminal loop of Rob can control the DNA binding mode, facilitating interconversion between the distinct DNA binding modes observed in MarA and Rob. In doing so, we provide detailed molecular insight into DNA binding and recognition by these proteins, which in turn is an important step towards the efficient design of anti-virulence agents that target these proteins.

中文翻译：

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转录因子MarA和Rob的N末端螺旋-螺旋-螺旋基序驱动DNA识别。

DNA结合蛋白在基因调节和细胞功能中起重要作用。转录因子MarA和Rob是AraC / XylS家族的两个同源成员，可调节多药耐药性。它们共享一个共同的DNA结合结构域，Rob拥有一个额外的C末端结构域，可以结合低分子量效应子。两种蛋白质都具有两个能够结合DNA的螺旋-转-螺旋（HTH）模体。然而，尽管MarA通过两个HTH基元与其启动子相互作用，但先前的研究表明，Rob通过单个HTH基元与DNA结合就足以紧密结合。在当前的工作中，我们执行两个转录因子与不同DNA序列结合的微秒级时标全原子模拟，以了解DNA识别和结合的决定因素。我们的模拟表征了DNA结合后动力学行为的序列特异性变化，展示了N末端HTH基序的Arg40在允许特异性紧密结合中的作用。最后，我们的模拟表明，Rob的酸性C末端环可以控制DNA结合模式，从而促进在MarA和Rob中观察到的不同DNA结合模式之间的相互转换。通过这样做，我们提供了对这些蛋白质与DNA结合和识别的详细分子了解，而这又是有效设计针对这些蛋白质的抗毒剂的重要一步。我们的模拟表明，Rob的酸性C末端环可以控制DNA结合模式，从而促进在MarA和Rob中观察到的不同DNA结合模式之间的相互转换。通过这样做，我们提供了对这些蛋白质与DNA结合和识别的详细分子了解，而这又是有效设计针对这些蛋白质的抗毒剂的重要一步。我们的模拟表明，Rob的酸性C末端环可以控制DNA结合模式，从而促进在MarA和Rob中观察到的不同DNA结合模式之间的相互转换。通过这样做，我们提供了对这些蛋白质与DNA结合和识别的详细分子了解，而这又是有效设计针对这些蛋白质的抗毒剂的重要一步。