title: Encoded protein from ycbR gene of enterohemorrhagic Escherichia coli O157:H7 associated with adherence to HEp-2 cells.
abstract: Adhesion is one of the significant virulence factors in enterohemorrhagic Escherichia coli (EHEC) O157:H7 pathogenesis. It is regulated by specific loci in the genome sequence. This study mainly focused on investigating the influence of ycbR gene and its encoded YCBR protein on the adhesion of EHEC O157:H7 to HEp-2 cells. In the first part, mutants of EHEC O157:H7 were constructed through TnphoA mutagenesis and assayed for adherent ability. Six mutant strains with lost adherence to HEp-2 cells were isolated and then sequenced using a primer that hybridized to phoA sequence downstream of the fusion joint. The sequencing results indicated that the gene of eae and ycbR, between the initiation codon and the -10 sequence of Z4182, yciI, ARAC-type regulator protein, and high-affinity gluconate transporter of EHEC were all possibly related to adhesion. Of the six genes, the ycbR gene was cloned to the pET28a vector to analyze its function further. Recombinant YCBR protein fused to a His tag (YCBR-His) was expressed under IPTG induction and purified by Ni-NTA column. The purified protein was subcutaneously injected to rabbits to prepare antisera. The results of an adherence assay in the presence of anti-YCBR-His antibodies indicated that antibodies blocked the adherence of EHEC O157:H7 to HEp-2 cells. These observations suggested that ycbR encoded a novel adherence-associated determinant of EHEC O157:H7, which could contribute to the adhesive capacity of the bacteria.
authors.affiliation: School of Chemical Engineering and Technology, Tianjin University, Tianjin, PR China.
authors.forename: Xiqian
authors.initials: X
authors.lastname: Tan
authors.affiliation: School of Chemical Engineering and Technology, Tianjin University, Tianjin, PR China.
authors.forename: Huazhi
authors.initials: H
authors.lastname: Xiao
authors.affiliation: School of Chemical Engineering and Technology, Tianjin University, Tianjin, PR China.
authors.forename: Ye
authors.initials: Y
authors.lastname: Han
authors.affiliation: School of Chemical Engineering and Technology, Tianjin University, Tianjin, PR China.
authors.forename: Xiaodi
authors.initials: X
authors.lastname: Hong
authors.affiliation: School of Chemical Engineering and Technology, Tianjin University, Tianjin, PR China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Cui
authors.affiliation: School of Chemical Engineering and Technology, Tianjin University, Tianjin, PR China. Electronic address: zzj@tju.edu.cn.
authors.forename: Zhijiang
authors.initials: Z
authors.lastname: Zhou
mesh_terms: D000818:Animals; D001422:Bacterial Adhesion; D004847:Epithelial Cells / Q000382:microbiology; D004927:Escherichia coli Infections / Q000382:microbiology*; D019453:Escherichia coli O157 / Q000235:genetics / Q000502:physiology*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D056945:Hep G2 Cells; D006801:Humans; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D011817:Rabbits
chemical_list: D029968:Escherichia coli Proteins; D018832:Molecular Chaperones; C000597920:YcbR protein, E coli
keywords: Adhesion; Enterohemorrhagic Escherichia coli O157:H7; ycbR