Khan A S, Schifferli D M
Department of Pathobiology, University of Pennsylvania School of Veterinary Medicine, Philadelphia 19104.
Infect Immun. 1994 Oct;62(10):4233-43. doi: 10.1128/iai.62.10.4233-4243.1994.
The 987P fimbriae produced by enterotoxigenic strains of Escherichia coli isolated from piglets mediate bacterial attachment to intestinal epithelial cells. These fimbriae consist essentially of a tight helical arrangement of one structural protein subunit encoded by fasA. Fimbriation and specific adhesion requires the expression of seven additional genes (fasB to fasH). In this study, we investigated whether FasA or another Fas protein, e.g., a potential minor fimbrial component, harbors the binding moiety for the pig 987P receptor glycoproteins. Fas proteins, specifically radiolabeled with an in vivo T7 expression system, were isolated from the periplasm and incubated with receptor-containing brush borders isolated from piglet intestinal epithelial cells. FasG bound best to brush borders, whereas no FasA adhered to them. Additional evidence that FasG, and not FasA, is the 987P adhesin was provided by ligand blotting inhibition assays indicating that FasG alone inhibited fimbrial binding to 987P receptors and that in the absence of FasG, other Fas proteins were not inhibitory. FasG was identified in purified fimbrial preparations with a specific anti-FasG antibody probe. Moreover, FasG was shown to be tightly associated with the fimbrial structure, since it was released only after disassembling fimbriae by heat and sodium dodecyl sulfate treatments. The primary structure of FasG, deduced from the DNA sequence, exhibited 19.1 to 24.4% similarity to FasA and large minor components and/or adhesins of other fimbriae. FasG is the first-described minor fimbrial subunit shown to be essential for both fimbrial biogenesis and specific adhesion.
从仔猪中分离出的产肠毒素大肠杆菌所产生的987P菌毛介导细菌与肠上皮细胞的附着。这些菌毛主要由fasA编码的一种结构蛋白亚基紧密螺旋排列组成。菌毛形成和特异性黏附需要另外七个基因（fasB至fasH）的表达。在本研究中，我们调查了FasA或另一种Fas蛋白（例如潜在的次要菌毛成分）是否含有猪987P受体糖蛋白的结合部分。利用体内T7表达系统进行特异性放射性标记的Fas蛋白从周质中分离出来，并与从仔猪肠上皮细胞分离出的含受体的刷状缘一起孵育。FasG与刷状缘结合最佳，而FasA不与之黏附。配体印迹抑制试验提供了额外证据，表明FasG而非FasA是987P黏附素，该试验表明单独的FasG可抑制菌毛与987P受体的结合，且在没有FasG的情况下，其他Fas蛋白无抑制作用。用特异性抗FasG抗体探针在纯化的菌毛制剂中鉴定出FasG。此外，FasG被证明与菌毛结构紧密相关，因为只有在通过加热和十二烷基硫酸钠处理使菌毛解体后它才会释放出来。从DNA序列推导的FasG一级结构与FasA以及其他菌毛的大的次要成分和/或黏附素具有19.1%至24.4%的相似性。FasG是首个被描述的对菌毛生物合成和特异性黏附都必不可少的次要菌毛亚基。
