细菌表面多糖抗原合成基因功能的系统研究

Bacterial surface polysaccharide antigen, such as O-antigen and K-antigen, exhibits one of the richest diversities in nature due to long-term pressure from the host immune system. It contributes major antigenic variability to the cell surface, and plays essential roles in the virulence and host adaptation of bacteria. Under the support by National Natural Science Foundation of Youth Science Fund project, the applicant’s work, including systematical comparative analysis of bacterial O-antigen gene clusters and structural analysis of polysaccharide antigens, has improved our understanding for the formation process of bacterial surface polysaccharide antigen diversity. In that project, a total of 19 SCI papers have been published up to now. Currently, there are still many gaps in the knowledge about the function of genes involved in the synthesis of O-antigen. Unique monosaccharides and oligosaccharides play important roles in the formation of bacterial surface polysaccharide diversity, and are beneficial for bacteria escaping from host immune recognition. Unique monosaccharides and oligosaccharides are also valuable in the filed of biopharmaceuticals. This project will identify several novel biosynthesis pathways for unique monosaccharides. It will also identify the function of several glycosyltransferases involved in the synthesis of surface antigens from important pathogens, and study the functional mechanism of these enzymes. The side chain modification of O-antigen has been reported to be related with the pathogenicity of Shigella. This project will systematically identify the genes responsible for the O-antigen modification of Escherichia coli and Salmonella, through the research methods of comparative genomics, molecular biology and etc, and elucidates their functional mechanism.

细菌表面多糖抗原（O抗原、K抗原等）在宿主免疫系统长期压力下形成了自然界中最丰富的生物多样性之一，是重要的免疫原性物质和致病因子。在青年科学基金项目中，申请人通过对大肠杆菌等细菌O抗原基因簇的系统比较分析和O抗原化学结构的解析，对于细菌表面多糖抗原多样性的形成方式有了较为深入的认识，相关结果已发表19篇SCI论文。目前，对于O抗原合成基因的功能研究尚存在较多空白。罕见单糖和寡糖在多糖抗原多样性的形成中起到重要作用，其有助于细菌逃避宿主细胞的免疫识别，并在生物制药等领域具有应用价值。本项目将鉴定一系列前期研究中新发现的罕见单糖生物合成途径；解析多个参与致病菌表面多糖抗原合成的糖基转移酶的功能，并解析其作用机制。O 抗原的侧链修饰已被报道可影响志贺氏菌的致病性和流行能力，本项目将利用比较基因组学等方法，系统发现和鉴定负责大肠杆菌和沙门氏菌O抗原侧链修饰的相关基因，并解析其侧链修饰机制。

细菌表面多糖抗原（O抗原、K抗原等）长期受到宿主免疫系统的选择压力，导致其形成了高度的多样性。细菌表面多糖抗原多样性是细菌血清型分型的基础。此外，它对于细菌的致病能力、以及在不同环境中生存的选择优势均具有重要意义。但目前，对于细菌表面多糖抗原合成基因的功能研究尚存在较多空白。本项目通过罕见单糖的生物合成途径、糖基转移酶的功能特异性等多个角度，系统研究了细菌表面多糖抗原合成基因行使功能的分子机制。. 本项目顺利完成了全部研究任务。具有罕见单糖的抗原结构更容易逃避宿主细胞的免疫识别，我们发现并鉴定了RhaNAc3NFo等四种罕见单糖的生物合成路径，相关工作也为利用生物技术体外合成罕见单糖奠定了重要基础。通过生物信息分析和生物化学实验，鉴定了大肠杆菌O3中糖基转移酶WfcD等9种糖基转移酶的功能；发现并鉴定了大肠杆菌O117、O107中的糖基转移酶WclY中与其功能特异性直接相关的位点；丰富了我们对于糖基转移酶功能特异性形成机制的认识。在痢疾志贺氏菌D13型、大肠杆菌O77型等6种细菌中，定位了O抗原侧链修饰基因，表明多糖抗原的侧链修饰是其多样性形成的重要机制。完整解析了沙门氏菌全部O抗原的多样性形成机制，发现大肠杆菌和沙门氏菌这两种重要致病菌的O抗原之间具有远高于预期的紧密亲缘关系，显示表面抗原多样性作为一种适应环境的重要方式在很久之前就存在于细菌中。首次定位了鲍曼不动杆菌表面多糖抗原基因簇的位点，并发现了一系列罕见单糖合成途径及新的糖基转移酶；建立了鲍曼不动杆菌分子生物学血清型分析技术，相比传统血清型鉴定方法具有快速、准确的优点，有望广泛应用于针对鲍曼不动杆菌的流行病调查和防控。. 项目累计在PNAS（影响因子9.4）、FEMS Microbiology Reviews（影响因子13.2）发表SCI论文14篇， 其中项目负责人为5篇论文的通讯作者、为6篇论文的第一（或并列第一）作者。

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