CS3抗原基因的表达及纤毛装配元件的研究

Study on the genetic determinant for expresson and assembly of CS3 fimbriae

在肠毒素大肠杆菌(ETEC)的已知定居因子中,CS3是临床分离株中最常见的抗原之一。为了研究CS3纤毛装配的基本元件,绘制了CS3亚基结构基因和辅助蛋白编码区的限制酶谱。通过亚克隆的亚基基因和不同辅助蛋白基因之间的互补性表达结果,确定了CS3纤毛装配所需要的辅助蛋白的DNA功能片段。微细胞分析结果显示,CS3基因的有效表达和纤毛装配至少需要6条蛋白多肽,分子量分别为(×10~3)15、17、24、27、48和90。除了15×10~3/17×10~3的蛋白多肽为CS3亚基外,其余的蛋白多肽参与CS3亚基的转运及纤毛的装配。根据以上结果初步确定了上述相关基因的相对位置。

Colonization factor antigens (CFAs) are important toxic and protective antigens of enterotox-igenic Escherichia coli (ETEC). Among the known CFAs, CS3 is a major fimbriae antigen. To study genes encoding CS3 subunit and auxiliary proteins which are responsible for assembly of CS3 fimbriae. the retriction map of CS3 genetic determinant was made. Bosed on the map, the structural gene of CS3 subunit and the region encoding auxiliary proteins were cloned respectively. Expression results of the recombinant plasmids in minicells showed that the genetic determinant of CS3 fimbriae encodes six polypeptides with the molecular weight 15 kD,17 kD,24 kD,48 kD and 90 kD,respectively. Western blotting and complemetary expression analysis among the different mutants showed 15 kD/17 kD proteins are CS3 subunit and it's precursor, the others may be necessary for assembly of CS3 fimbriae. The relative location of various genes of CS3 genetic determinant was determined.