title: Characterization of the role in adherence of Mycoplasma hyorhinis variable lipoproteins containing different repeat unit copy numbers.
abstract: Mycoplasma hyorhinis (M. hyorhinis) is an important pathogen of pigs. In previous studies, the variable lipoprotein (Vlp) family has been shown to play a role in mediating M. hyorhinis cytoadhesion. Herein, we performed several experiments to study the function of each Vlp family member in detail, especially examining the cytoadhesion functional domain and how the repeat unit copy number impacts on function. Recombinant proteins rVlpII, composed of region II from all seven Vlp members; rVlpIII, composed of repeat peptides from region III of all of Vlp members; as well as a series of recombinant rVlp proteins for each member containing different repeat unit copy numbers were constructed. All of the proteins were expressed in Escherichia coli and purified by affinity chromatography. The recombinant proteins, as well as seven keyhole limpet hemocyanin-conjugated Vlp peptides containing two copies of the repeat unit, were analyzed for their adherence to swine tracheal epithelial cells using a microtiter plate adherence assay. Both rVlpII and rVlpIII proteins were able to bind to cell membrane proteins. Among the repeat unit peptides, only PepVlpB and PepVlpG were able to bind to cell membrane proteins. All of the Vlp members had cytoadhesion capability. The adhesion abilities of the proteins containing 0 or 3 copies of the repeat unit were stronger than those of the proteins containing 12 copies. For rVlpA, rVlpB, rVlpD, rVlpF and rVlpG, the proteins containing no copies bound stronger than the proteins containing 3 copies. In contrast, the adherence of rVlpC3 was stronger than that of rVlpC0. There was no significant difference between the adherence of rVlpE3 and that of rVlpE0. Our results suggest that the major cytoadhesion sites of Vlps are mainly contained in region II, the function of which would be blocked by region III when region III is longer.
authors.affiliation: Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, National Center for Engineering Research of Veterinary Bio-products, Nanjing 210014, China.
authors.forename: Qiyan
authors.initials: Q
authors.lastname: Xiong
authors.affiliation: Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, National Center for Engineering Research of Veterinary Bio-products, Nanjing 210014, China; Shanxi Agricultural University, Taigu, Shanxi 030801, China.
authors.forename: Bixiong
authors.initials: B
authors.lastname: Zhang
authors.affiliation: Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, National Center for Engineering Research of Veterinary Bio-products, Nanjing 210014, China.
authors.forename: Jia
authors.initials: J
authors.lastname: Wang
authors.affiliation: Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, National Center for Engineering Research of Veterinary Bio-products, Nanjing 210014, China.
authors.forename: Bo
authors.initials: B
authors.lastname: Ni
authors.affiliation: Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, National Center for Engineering Research of Veterinary Bio-products, Nanjing 210014, China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Ji
authors.affiliation: Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, National Center for Engineering Research of Veterinary Bio-products, Nanjing 210014, China.
authors.forename: Yanna
authors.initials: Y
authors.lastname: Wei
authors.affiliation: Division of Animal Infectious Diseases, State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China.
authors.forename: Shaobo
authors.initials: S
authors.lastname: Xiao
authors.affiliation: Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, National Center for Engineering Research of Veterinary Bio-products, Nanjing 210014, China.
authors.forename: Zhixin
authors.initials: Z
authors.lastname: Feng
authors.affiliation: Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, National Center for Engineering Research of Veterinary Bio-products, Nanjing 210014, China.
authors.forename: Maojun
authors.initials: M
authors.lastname: Liu
authors.affiliation: Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Veterinary Biological Engineering and Technology, Ministry of Agriculture, National Center for Engineering Research of Veterinary Bio-products, Nanjing 210014, China; Jiangsu Collaborative Innovation Center for Meat Production, Processing and Quality control, Nanjing 210014, China. Electronic address: gqshaojaas@163.com.
authors.forename: Guoqing
authors.initials: G
authors.lastname: Shao
mesh_terms: D000818:Animals; D001422:Bacterial Adhesion / Q000502:physiology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000502:physiology; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D008074:Lipoproteins / Q000378:metabolism*; D045790:Mycoplasma hyorhinis / Q000502:physiology*; D000072417:Protein Domains; D011994:Recombinant Proteins / Q000235:genetics; D020545:Respiratory Mucosa / Q000166:cytology; D013552:Swine
chemical_list: D001426:Bacterial Proteins; D008074:Lipoproteins; D011994:Recombinant Proteins
keywords: Adhesion; Copy number; Mycoplasma hyorhinis; Variable lipoprotein