Lulu Chen1, Xin Li2, Xuedong Zhou3, Jumei Zeng4, Zhi Ren3, Lei Lei5, Di Kang3, Keke Zhang2, Jing Zou1, Yuqing Li6. 1. State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, PR China; Department of Pediatric Dentistry, West China Hospital of Stomatology, Sichuan University, Chengdu, PR China. 2. State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, PR China. 3. State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, PR China; Department of Endodontics Dentistry, West China Hospital of Stomatology, Sichuan University, Chengdu, PR China. 4. Department of Infectious Diseases, Boston Children's Hospital, Harvard Medical School, Boston, Massachusetts. 5. State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, PR China; Department of Preventive Dentistry, West China Hospital of Stomatology, Sichuan University, Chengdu, PR China. 6. State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, PR China. Electronic address: liyuqing@scu.edu.cn.
Abstract
INTRODUCTION: Enterococcus faecalis is correlated with oral diseases including recurrent root canal treatment failure because of its biofilm formation ability and various virulence factors. Cyclic di-AMP (c-di-AMP) is an omnipresent second messenger involved in many crucial cellular physiological processes, including biofilm formation. ST056083 is a small molecule working as an inhibitor of the c-di-AMP synthetase DNA integrity scanning protein (DisA) in vitro. In this study, the impact of ST056083 on E. faecalis DisA activity, bacterial growth, and biofilm formation was tested. METHODS: The binding affinity between the protein and ligand was evaluated using the Amber score, and the binding mode was analyzed and visualized using UCSF Chimera (Resource for Biocomputing, Visualization, and Informatics, University of California, San Francisco, San Francisco, CA). The effect of ST056083 on E. faecalis DisA was evaluated using the coralyne assay. The effect of ST056083 on E. faecalis biofilm formation was determined by the biofilm quantification assay, scanning electron microscopic examination, and 3-dimensional confocal laser scanning microscopic assay. The effect of ST056083 on E. faecalis exopolysaccharide synthesis was measured by the anthrone-sulfuric method. RESULTS: We expressed and purified E. faecalis DisA in vitro and confirmed the inhibitory effect of ST056083 on its biological activity. In addition, we showed the inhibitory effect of ST056083 on E. faecalis growth, biofilm formation, and exopolysaccharide synthesis. CONCLUSIONS: Our findings enhance the understanding of the physiological role of c-di-AMP in E. faecalis and represent a preliminary study on the ST056083 inhibitory effect and mechanism.
INTRODUCTION:Enterococcus faecalis is correlated with oral diseases including recurrent root canal treatment failure because of its biofilm formation ability and various virulence factors. Cyclic di-AMP (c-di-AMP) is an omnipresent second messenger involved in many crucial cellular physiological processes, including biofilm formation. ST056083 is a small molecule working as an inhibitor of the c-di-AMP synthetase DNA integrity scanning protein (DisA) in vitro. In this study, the impact of ST056083 on E. faecalis DisA activity, bacterial growth, and biofilm formation was tested. METHODS: The binding affinity between the protein and ligand was evaluated using the Amber score, and the binding mode was analyzed and visualized using UCSF Chimera (Resource for Biocomputing, Visualization, and Informatics, University of California, San Francisco, San Francisco, CA). The effect of ST056083 on E. faecalis DisA was evaluated using the coralyne assay. The effect of ST056083 on E. faecalis biofilm formation was determined by the biofilm quantification assay, scanning electron microscopic examination, and 3-dimensional confocal laser scanning microscopic assay. The effect of ST056083 on E. faecalisexopolysaccharide synthesis was measured by the anthrone-sulfuric method. RESULTS: We expressed and purified E. faecalis DisA in vitro and confirmed the inhibitory effect of ST056083 on its biological activity. In addition, we showed the inhibitory effect of ST056083 on E. faecalis growth, biofilm formation, and exopolysaccharide synthesis. CONCLUSIONS: Our findings enhance the understanding of the physiological role of c-di-AMP in E. faecalis and represent a preliminary study on the ST056083 inhibitory effect and mechanism.
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