Structural characterization of the enterococcal polysaccharide antigen and analysis of its contribution to cell growth, division and antibiotic resist

拟议的项目重点是对所有肠球菌产生的一种表面鼠李多糖进行功能分析，这是正常细胞生长、分裂、抗药性和致病所必需的。这种肠球菌多糖抗原(EPA)由两个基因簇编码：(I)18个极其保守的基因(EPAA-EPAR)编码一个核心合成机制；(Ii)10-20个基因在不同菌株之间变异，负责修饰多糖骨架。我们已经构建了完全缺失EPA可变区的突变体，并表明EPA的修饰(但不是其核心结构)对于这种聚合物的生物活性是必不可少的。我们建议研究EPA的结构性质，剖析这种聚合物及其修饰对细菌生理和抗生素耐药性的具体贡献。我们将采取多学科的方法来探索EPA的结构，了解EPA如何控制参与细胞壁组装的表面蛋白的分布和活性以及对抗生素的耐药性。该项目将涉及最先进的方法，包括超分辨率显微镜(荧光和电子显微镜)、结构糖生物学(核磁共振、质谱学)和研究细菌生理学和抗菌素耐药性的传统方法。候选人将与监督实验室的成员合作，研究细菌细胞表面组装和宿主与病原体的相互作用。

The proposed project is focused on the functional analysis of a surfacerhamnopolysaccharide produced by all enterococci, required for normal cell growth,division, resistance to antibiotics and pathogenesis. This enterococcal polysaccharideantigen (EPA) is encoded by two gene clusters: (i) 18 genes extremely conserved (epaA-epaR) encoding a core synthetic machinery and (ii) 10-20 genes variable from one strain toanother, responsible for the decoration of the polysaccharide backbone. We have built amutant with a complete deletion of the epa variable region and shown that the decoration ofEPA (but not its core structure) is essential for the biological activity of this polymer. Wepropose investigate the structural properties of Epa and dissect the specific contribution ofthis polymer and its decoration to bacterial physiology and antibiotic resistance.We will take a multidisciplinary approach to explore EPA structure and to understand howEPA controls the distribution and activity of surface proteins involved in cell wall assemblyand resistance to antibiotics. The project will involve state-of-the-art methodologiesincluding super-resolution microscopy (fluorescence and electron microscopy), structuralglycobiology (NMR, mass spectrometry) and conventional approaches to study bacterialphysiology and antimicrobial resistance. The candidate will work with members of thesupervisors' laboratories studying bacterial cell surface assembly and host-pathogeninteractions.