新型Kdo2-lipidA合成菌株构建及其免疫生化活性研究  

Construction of Escherichia coli Mutant Producing Novel Kdo_2-Lipid A and Study of Its Immunological and Biochemical Activities

作　　者：王碧雯 李颜颜[1,2] 王小元 WANG Biwen;LI Yanyan;WANG Xiaoyuan(State Key Laboratory of Food Science and Technology,Jiangnan University,Wuxi 214122,China;International Joint Laboratory on Food Safety,Jiangnan University,Wuxi 214122,China)

机构地区：[1]食品科学与技术国家重点实验室,江南大学,江苏无锡214122 [2]食品安全国际联合实验室,江南大学,江苏无锡214122

出　　处：《食品与生物技术学报》2018年第5期479-486,共8页Journal of Food Science and Biotechnology

基　　金：国家重点研发计划“食品安全关键技术研发”专项(2017YFC1600102); 国家轻工技术与工程一流学科自主课题(LITE2018-10)

摘　　要：脂多糖(LPS)是革兰氏阴性细菌外膜的主要组成部分,由疏水的Kdo2-lipid A和亲水的多糖长链组成,能激活先天性免疫系统。Kdo2-lipid A是LPS的生物活性中心,能被开发为新型疫苗佐剂。本研究在前期菌株基础上敲除与LPS庚糖合成相关waaC-waaF基因,构建了直接合成特殊结构Kdo2-lipid A、不含多糖长链的突变株BW003。通过薄层层析和电喷雾电离质谱技术鉴定了BW003合成的Kdo2-lipid A结构;HEK-Blue h TLR4细胞和THP-1细胞刺激实验证实突变株及其合成的Kdo2-lipid A的细胞毒性均明显减弱;生化表型分析结果表明BW003较野生型呈现出更高的外膜渗透性、抗生素敏感性、疏水性及自凝集能力。本研究为后续新型疫苗佐剂开发和产业化利用奠定了理论基础。Lipopolysaccharides（LPS） is the major component of the outer membrane of Gram-negative bacteria. It＇s composed of a hydrophobic Kdo2-lipid A and a hydrophilic polysaccharide chain,and can activate the innate immune system. Kdo2-lipid A is the bioactive centers of LPS,which has the potential to be developed as vaccines or vaccine adjuvants. In this study,we constructed a novel Kdo2-lipid A producing mutant BW003 by deleting waaC-waaF genes in the chromosome of a laboratory Escherichia coli strain. The Kdo2-lipid A structure of BW003 was confirmed by TLC and ESI/MS. Compared with wild type E. coli W3110 and LPS,BW003 and Kdo2-lipid A showed lower cytotoxicity when stimulating HEK-Blue h TLR4 cells and THP-1 cells.In addition,BW003 presented higher outer membrane permeability,sensitivity to antibiotics,hydrophobicity and auto-aggregation ability,which is safe and efficient for large-scale industrial fermentation and vaccine adjuvants development.

关 键 词：脂多糖 Kdo2-lipid A 免疫生化活性 waaC-waaF基因 疫苗佐剂 

分 类 号：Q939.91[生物学—微生物学]