MOLECULAR CHARACTERIZATION OF A CARBOXY-TERMINAL EUKARYOTIC-CELL-BINDING DOMAIN OF INTIMIN FROM ENTEROPATHOGENIC ESCHERICHIA-COLI
MOLECULAR CHARACTERIZATION OF A CARBOXY-TERMINAL EUKARYOTIC-CELL-BINDING DOMAIN OF INTIMIN FROM ENTEROPATHOGENIC ESCHERICHIA-COLI
复制标题
DOI：
10.1128/iai.63.11.4323-4328.1995
复制
发表时间：
1995-11-01
影响因子：
3.1
通讯作者：
DOUGAN, G
中科院分区：
文献类型：
--
作者：
FRANKEL, G;CANDY, DCA;DOUGAN, G
研究了来自肠致病性大肠杆菌 O127 (EPEC) 的 intimin (Int) 多肽的真核细胞结合结构域，将 Int (Int(EPEC280)) 的羧基末端 280 个氨基酸结构域和来自假结核耶尔森氏菌 (Inv(YP280)) 的 Int 同源入侵素 (Inv) 的衍生物融合到E.调用麦芽糖结合蛋白(MBP)，表达并纯化。通过在荧光激活细胞分选仪分析中使用纯化的融合蛋白或通过使用涂有纯化融合物的荧光 Covaspheres 进行监测，有效介导与 HEp-2 细胞结合的最小 MBP-Int(EPEC) 融合蛋白含有 Int 的羧基末端 150 个氨基酸。用 Ser (Int(EPEC280CS)) 替换 Cys-937 破坏了 Int(EPEC280) 的细胞结合活性。涂有 MBP-Int (EPEC280) 的 Covaspheres 与 HEp-2 细胞微绒毛相关，但未能诱导肌动蛋白在结合颗粒下方积聚或细胞在涂层塑料表面上铺展，MBP-Int (EPEC280) 但 MBP、MBP-Int (EPEC280CS) 或 MBP-Inv (YP280) 不会抑制 EPEC 进入 HEp-2 细胞。
A eukaryotic cell-binding domain from the intimin (Int) polypeptide of enteropathogenic Escherichia coli O127 (EPEC) was investigated, Derivatives of the carboxy-terminal 280-amino-acid domains of Int (Int(EPEC280)) and the Int homolog invasin (Inv) from Yersinia pseudotuberculosis (Inv(YP280)) were fused to the E. call maltose-binding protein (MBP), expressed, and purified. The smallest MBP-Int(EPEC) fusion protein that efficiently mediated binding to HEp-2 cells, monitored by using purified fusion proteins in fluorescence activated cell sorter analysis or by using fluorescent Covaspheres coated with purified fusions, contained the carboxy-terminal 150 amino acids of Int. Replacement of Cys-937 with Ser (Int(EPEC280CS)) destroyed the cell-binding activity of Int(EPEC280). Covaspheres coated with MBP-Int(EPEC280) were associated with HEp-2 cell microvilli but failed to induce actin accumulation underneath bound particles or cell spreading on coated plastic surfaces, MBP-Int(EPEC280), but not MBP, MBP-Int(EPEC280CS), or MBP-Inv(YP280), inhibited EPEC entry into HEp-2 cells.