The recognition and binding of specific DNA promotors by transcription factors (TF) is a key aspect in gene regulation. Rob and MarA are two homologous TF, members of the AraC/XylS family, involved in the regulation of multiple genes involved in multidrug resistance. They share a common DNA binding domain of around 100 residues arranged in two helix-turn-helix (HTH) motifs, common to all members of this family. This domain presents 51% sequence identity and 71% sequence similarity between MarA and Rob, allowing for some overlap between the DNA promotors specific to these TF, although with different activation efficiencies. While MarA consist in only this domain, Rob possess an extra C-terminal domain. X-ray structures however shows that these TF establish different interactions with their promotors, with MarA being able to interact with mar promotor via both HTH motifs while Rob can interact with micF promotor via only one of the HTH motifs. In the present work, we performed all-atoms simulations on the microsecond time scale, in order to understand the molecular details of Rob and MarA binding to different DNA sequences. Our simulations have explored the intrinsic flexibility of both free TF and their DNA promotors, and provided useful insights into the sequence-specific changes in dynamical behaviour upon DNA binding by the two TF. It also showcases the role of Arg40, present in the HTH-motif, in allowing for specific tight binding. They also clearly demonstrate the role of an acidic C-terminal loop of Rob that can control the DNA binding mode, explaining the distinct DNA binding modes observed in MarA and Rob. With the analysis of our simulations, we provide detailed molecular insight into DNA binding and recognition by both MarA and Rob, which in turn is an important step towards the efficient design of anti-virulence agents that target these proteins.

Insights into the DNA recognition by Mara and Rob

ACS Spring Meeting 2021

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