Mechanism Of Reduced Vancomycin Susceptibility Conferred By WalK Mutation In Staphylococcus Aureus

Cell wall-active antibiotics are typically bactericidal, in contrast to inhibitors of protein synthesis, which are frequently bacteriostatic. Thus, killing often follows inhibition of peptidoglycan biosynthesis, and this is often accompanied by cellular lysis. Unfortunately, bacteria have not remained susceptible to cell wall-active antibiotics, and resistance to these agents is common with the adaptive evolution of bacteria. Penicillin resistance due to P-lactamase production had become common by the end of the nineteen fifties. The P-lactamase-resistant methicillin was introduced in the early nineteen sixties. Methicillin-resistant Staphylococcus aureus has been common in hospital-acquired infections for a couple of decades, and exhibits resistance to many different classes of antibiotics. Vancomycin intermediate-resistant Staphylococcus aureus was firstly reported in 1997 and then fully vancomycin-resistant S. aureus was described in the US. The clinical data suggest that pan-resistant 5. aureus strains certainly appear to be a possibility. This work focuses on the investigation of the molecular mechanisms of vancomycin-intermediate resistance, and aims to provide a theoretical basis for novle drug targets.Point mutations are often associated with antibiotic resistance in clinical bacterial isolates, but the molecular mechanism underlying the emergence of the resistance remains largely unknown. By antibiotic screening, we generated a vancomycin intermediate-resistant strain from S. aureus strain MW2. Genetic changes in the mutant were identified by genome sequencing and G223D mutation was found in WalK, which is a sensor protein of the two-component system walKR. By introducing the walK (G223D) mutation into the wild-type strain MW2, we observed that bacterial cells with the point mutation exhibited decreased expression of genes associated with cell wall metabolism, decreased autolytic activity, thickened cell wall, and reduced vancomycin susceptibility. Phosphorylation assay showed that WalK (G223D) exhibited reduced autophosphorylation, which led to reduced phosphorylation of WalR. Electrophoretic mobility shift assay indicated that WalK (G223D)-phosphorylated WalR had a reduced binding capacity to atlA promoter. In conclusion, WalK (G223D) exhibited reduced autophosphorylation, which led to reduced phosphorylation of WalR. Reduced binding capacity of WalK (G223D)-phosphorylated WalR to atlA promoter decreased the expression of atlA, which led to decreased cell wall metabolism. Subsequently, the mutant presents decreased autolytic activity, thickened cell wall, and reduced vancomycin susceptibility.