P27 (MBOV_RS03440) is a novel fibronectin binding adhesin of Mycoplasma bovis
P27 (MBOV_RS03440) is a novel fibronectin binding adhesin of Mycoplasma bovis
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DOI：
10.1016/j.ijmm.2018.07.006
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发表时间：
2018-10-01
影响因子：
4.1
通讯作者：
Guo, Aizhen
中科院分区：
文献类型：
--
作者：
Chen, Xi;Huang, Jing;Guo, Aizhen
牛支原体是引起牛呼吸道疾病的主要病原体之一，它与呼吸道上皮细胞结合，导致严重的肺炎和组织损伤。本研究旨在鉴定27 kDa牛分枝杆菌脂蛋白的黏附功能，该蛋白由MBOV_RS03440基因编码，命名为P27。克隆并高效表达重组P27(RP27)抗体。Western印迹和流式细胞仪检测证实P27是一种表面定位蛋白，而ELISA法证实它是一种免疫原性蛋白。共聚焦免疫荧光显微镜显示rP27以剂量依赖的方式与胚胎牛肺(EBL)细胞结合。此外，抗rP27抗血清可抑制牛分枝杆菌与EBL细胞的黏附，表明P27具有与EBL细胞结合的特异性。RP27与EBL细胞的附着是由细胞外基质成分纤维连接蛋白(FN)介导的。用配基免疫印迹法、ELISA法和生物层析干涉法对rP27与FN之间的相互作用进行了定性和定量的监测。综上所述，这些结果表明P27是一种新的结合牛分枝杆菌的免疫原性黏附蛋白，从而有助于进一步了解牛分枝杆菌的分子发病机制。
Mycoplasma bovis, one of the major pathogens of bovine respiratory disease, binds to respiratory epithelial cells resulting in severe pneumonia and tissue damage. This study was designed to identify the adhesive function of a putative 27-kDa M. bovis lipoprotein, encoded by the gene MBOV_RS03440 and designated as P27. The gene was cloned and overexpressed to produce antibodies against the recombinant P27 (rP27). The western blot and flow cytometry assay confirmed P27 to be a surface-localized protein, while ELISA confirmed it to be an immunogenic protein. Confocal immunofluorescence microscopy demonstrated that rP27 bound to embryonic bovine lung (EBL) cell monolayers in a dose-dependent manner. Furthermore, anti-rP27 antiserum inhibited the attachment of M. bovis to EBL cells demonstrating the binding specificity of P27 to EBL cells. The attachment of rP27 to EBL cells was mediated by fibronectin (Fn), an extracellular matrix component. The interaction between rP27 and Fn was qualitatively and quantitatively monitored by ligand immunoblot assay, ELISA, and biolayer interferometry. Collectively, these results indicate that P27 is a novel En-binding, immunogenic adhesive protein of M. bovis, thereby contributing to the further understanding of the molecular pathogenesis of M. bovis.