TcaR-ssDNA complex crystal structure reveals new DNA binding mechanism of the MarR family proteins.

The teicoplanin-associated locus regulator (TcaR) regulates gene expression of proteins on the intercellular adhesion (ica) locus involved in staphylococci poly-N-acetylglucosamine biosynthesis. The absence of TcaR increases poly-N-acetylglucosamine production and promotes biofilm formation. Until recently, the mechanism of multiple antibiotic resistance regulator family protein members, such as TcaR, was restricted to binding double-stranded DNA. However, we recently found that TcaR strongly interacts with single-stranded DNA, which is a new role for this family of proteins. In this study, we report Staphylococcus epidermidis TcaR-single-stranded DNA complex structures. Our model suggests that TcaR and single-stranded DNA form a 61-symmetry polymer composed of TcaR dimers with single-stranded DNA that wraps outside the polymer and 12 nt per TcaR dimer. Single-stranded DNA binding to TcaR involves a large conformational change at the DNA binding lobe. Several point mutations involving the single-stranded DNA binding surface validate interactions between single-stranded DNA and TcaR. Our results extend the novel role of multiple antibiotic resistance regulator family proteins in staphylococci.

Structural analysis of the antibiotic-recognition mechanism of MarR proteins.

Staphylococci cause a wide range of diseases in humans and animals, and the proteins of the multiple antibiotic-resistance repressor (MarR) family in staphylococci function as regulators of protein expression and confer resistance to multiple antibiotics. Diverse mechanisms such as biofilm formation, drug transport, drug modification etc. are associated with this resistance. In this study, crystal structures of the Staphylococcus aureus MarR homologue SAR2349 and its complex with salicylate and the aminoglycoside antibiotic kanamycin have been determined. The structure of SAR2349 shows for the first time that a MarR protein can interact directly with different classes of ligands simultaneously and highlights the importance and versatility of regulatory systems in bacterial antibiotic resistance. The three-dimensional structures of TcaR from S. epidermidis in complexes with chloramphenicol and with the aminoglycoside antibiotic streptomycin were also investigated. The crystal structures of the TcaR and SAR2349 complexes illustrate a general antibiotic-regulated resistance mechanism that may extend to other MarR proteins. To reveal the regulatory mechanism of the MarR proteins, the protein structures of this family were further compared and three possible mechanisms of regulation are proposed. These results are of general interest because they reveal a remarkably broad spectrum of ligand-binding modes of the multifunctional MarR proteins. This finding provides further understanding of antimicrobial resistance mechanisms in pathogens and strategies to develop new therapies against pathogens.