Structural insights into DNA binding domain of vancomycin-resistance-associated response regulator in complex with its promoter DNA from Staphylococcus aureus.

In Staphylococcus aureus, vancomycin-resistance-associated response regulator (VraR) is a part of the VraSR two-component system, which is responsible for activating a cell wall-stress stimulon in response to an antibiotic that inhibits cell wall formation. Two VraR-binding sites have been identified: R1 and R2 in the vraSR operon control region. However, the binding of VraR to a promoter DNA enhancing downstream gene expression remains unclear. VraR contains a conserved N-terminal receiver domain (VraR _N ) connected to a C-terminal DNA binding domain (VraR _C ) with a flexible linker. Here, we present the crystal structure of VraR _C alone and in complex with R1-DNA in 1.87- and 2.0-Å resolution, respectively. VraR _C consisting of four α-helices forms a dimer when interacting with R1-DNA. In the VraR _C -DNA complex structure, Mg ²⁺ ion is bound to Asp194. Biolayer interferometry experiments revealed that the addition of Mg ²⁺ to VraR _C enhanced its DNA binding affinity by eightfold. In addition, interpretation of NMR titrations between VraR _C with R1- and R2-DNA revealed the essential residues that might play a crucial role in interacting with DNA of the vraSR operon. The structural information could help in designing and screening potential therapeutics/inhibitors to deal with antibiotic-resistant S. aureus via targeting VraR.
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