The MarR family transcriptional regulators serve as sensors of changing environments, allowing pathogenic bacteria to survive and persist in a dynamic environment [39] . However, up to now, the knowledge of MarR family protein-nucleic acid interaction has been focused on dsDNA and the MarR family protein-ssDNA interaction ability as well as their contribution to the multiple functions of TcaR is yet to be discovered. Better understanding of these interactions not only will benefit the understanding of many biological mechanisms but also is expected to provide a concept for designing a new therapy for Streptococci. In this work, we present the first attempt to investigate the TcaR-ssDNA interaction. The information of TcaR-ssDNA binding mode and the minimal binding length that we obtained from EMSA analysis and Biacore will be helpful for us to obtain TcaR-ssDNA complexed structure successfully. Moreover, we used in vitro replication assay and plaque assay to elucidate the specific biological role of the ssDNA binding ability of TcaR. Such observations may help us understand the mechanism of antibiotic resistance in the MarR family regulators.