Preliminary Crystallographic Study Of Carious Streptococcus Mutants Proteins

Objective:Dental caries (tooth decay) is a kind of common oral disease worldwide, it is a serious disease affect human health and quality of life. Streptococcus mutans (S. mutans) is the leading cause of dental caries. Using the structural genomics research methods, can better understand the gene complexity and specificity of S. mutans. Master how the S. mutans absorb nutrition and defense the host to adapt the oral environment. The genome analysis lays a foundation for prevention of dental caries and related drug design in the future.Methods:Protein crystallography is the tool of the structure genomics, it can analysis the three-dimensional protein structure and further apply in structure-based drug design study. Structural biology lab in Peking University has successfully constructed a high-throughput structural genomics laboratorial platform. The whole experimental flowchart, target genes selecting, high-throughput automatic gene cloning and expression, high-throughput protein purification, crystal screening, X-ray diffraction data collection system and structure determination, built a good platform for the research on the structural genomics of S. mutans.Results:This thesis mainly focuses on the purification and crystallization of the two proteins of those S. mutans genes selected. They are SMU.2055, SMU.1254. They are all cloned into pET28a. SMU.2055 and SMU.1254 expressed from the E. coli strain BL21 (DE3) and had good solubility. They were purified to homogeneity in a two-step procedure of Ni2+chelating and size exclusion chromatography. Then preliminary crystals were obtained either via sitting-drop vapor diffusion method. After optimization, some of them gave crystals suitable for X-ray diffraction and data collection. Among them, the data of both native and Se-Met protein of SMU.2055 have been collected and the structure is solved (PDB:3LD2). The data of native and heavy-atom soaking protein of SMU.1254 have been collected and the structure is undergoing.Conclusion:Structure genomics and protein crystallography research of carious S. mutans SMU.2055 protein and SMU.1254 protein, at the atomic level understand the structures and functions of those proteins of S. mutans. This study has built a good foundation for further understand and research of S. mutans biological metabolism mechanism and biology function in the dental caries.