Konkel M E, Garvis S G, Tipton S L, Anderson D E, Cieplak W
Department of Microbiology, Washington State University, Pullman 99164-4233, USA.
Mol Microbiol. 1997 Jun;24(5):953-63. doi: 10.1046/j.1365-2958.1997.4031771.x.
Campylobacter jejuni, a Gram-negative bacterium, is a common cause of gastrointestinal disease. By analogy with other enteric pathogens such as Salmonella and Shigella, the ability of C. jejuni to bind to host cells is thought to be essential in the pathogenesis of enteritis. Scanning electron microscopy of infected INT407 cells suggested that C. jejuni bound to a component of the extracellular matrix. Binding assays using immobilized extracellular matrix proteins and soluble fibronectin showed specific and saturable binding of fibronectin to C. jejuni. Ligand immunoblot assays using 125I-labelled fibronectin revealed specific binding to an outer membrane protein with an apparent molecular mass of 37 kDa. A rabbit antiserum, raised against the gel-purified protein, reacted with a 37 kDa protein in all C. jejuni isolates (n = 15) as tested by immunoblot analysis. Antibodies present in convalescent serum from C. jejuni-infected individuals also recognized a 37 kDa protein. The gene encoding the immunoreactive 37kDa protein was cloned and sequenced. Sequencing of overlapping DNA fragments revealed an open reading frame (ORF) that encodes a protein of 326 amino acids with a calculated molecular mass of 36872Da. The deduced amino acid sequence of the ORF exhibited 52% similarity and 28% identity to the root adhesin protein from Pseudomonas fluorescens. Isogenic C. jejuni mutants which lack the 37 kDa outer membrane protein, which we have termed CadF, displayed significantly reduced binding to fibronectin. Biotinylated fibronectin bound to a protein with an apparent molecular mass of 37 kDa in the outer membrane protein extracts from wild-type C. jejuni as judged by ligand-binding blots. These results indicate that the binding of C. jejuni to fibronectin is mediated by the 37 kDa outer membrane protein which is conserved among C. jejuni isolates.
空肠弯曲菌是一种革兰氏阴性菌,是胃肠道疾病的常见病因。与其他肠道病原体如沙门氏菌和志贺氏菌类似,空肠弯曲菌与宿主细胞结合的能力被认为在肠炎发病机制中至关重要。对感染的INT407细胞进行扫描电子显微镜观察表明,空肠弯曲菌与细胞外基质的一种成分结合。使用固定化细胞外基质蛋白和可溶性纤连蛋白的结合试验显示,纤连蛋白与空肠弯曲菌有特异性和饱和性结合。使用125I标记的纤连蛋白进行的配体免疫印迹试验显示,与一种表观分子量为37 kDa的外膜蛋白有特异性结合。用凝胶纯化蛋白制备的兔抗血清,通过免疫印迹分析检测,与所有空肠弯曲菌分离株(n = 15)中的一种37 kDa蛋白发生反应。空肠弯曲菌感染个体恢复期血清中的抗体也识别一种37 kDa蛋白。编码具有免疫反应性的37 kDa蛋白的基因被克隆并测序。重叠DNA片段的测序揭示了一个开放阅读框(ORF),其编码一个由326个氨基酸组成的蛋白,计算分子量为36872 Da。该ORF推导的氨基酸序列与荧光假单胞菌的根粘附素蛋白有52%的相似性和28%的同一性。缺乏我们称为CadF的37 kDa外膜蛋白的空肠弯曲菌同基因突变体,与纤连蛋白的结合显著减少。通过配体结合印迹判断,生物素化的纤连蛋白与野生型空肠弯曲菌外膜蛋白提取物中一种表观分子量为37 kDa的蛋白结合。这些结果表明,空肠弯曲菌与纤连蛋白的结合是由一种在空肠弯曲菌分离株中保守的37 kDa外膜蛋白介导的。