Abstract
Bacterial cell wall (CW) and extracellular (EC) proteins are often involved in interactions with extracellular matrix (ECM) proteins such as laminin (LN) and fibronectin (FN), which play important roles in adhesion and invasion. In this study, an efficient method combining proteomic analysis and Far-Western blotting assays was developed to screen directly for bacterial surface proteins with LN- and FN-binding capacity. With this approach, fifteen potential LN-binding proteins and five potential FN-binding proteins were identified from Streptococcus suis serotype 2 (SS2) CW and EC proteins. Nine newly identified proteins, including oligopeptide-binding protein OppA precursor (OppA), elongation factor Tu (EF-Tu), enolase, lactate dehydrogenase (LDH), fructose-bisphosphate aldolase (FBA), 3-ketoacyl-ACP reductase (KAR), Gly ceraldehyde-3-phosphate dehydrogenase (GAPDH), Inosine 5′-monophosphate dehydrogenase (IMPDH), and amino acid ABC transporter permease (ABC) were cloned, expressed, purified and further confirmed by Far-Western blotting and ELISA. Five proteins (OppA, EF-Tu, enolase, LDH, and FBA) exhibited specifically binding activity to both human LN and human FN. Furthermore, seven important recombinant proteins were selected and identified to have the ability to bind Hep-2 cells by the indirect immunofluorescent assay. In addition, four recombinant proteins, and their corresponding polyclonal antibodies, were observed to decrease SS2 adhesion to Hep-2 cells, which indicates that these proteins contribute to the adherence of SS2 to host cell surface. Collectively, these results show that the approach described here represents a useful tool for investigating the host-pathogen interactions.
Highlights
Streptococcus suis (SS) is an important pathogen in swine and human that causes septicemia, meningitis, arthritis, and pneumonia (Gottschalk and Segura, 2000)
By using the same approach, suis serotype 2 (SS2) cell wall (CW) and EC protein spots in the 2-DE gels were stained with the Coomassie brilliant blue G250 (Figures 1E,G) and five potential SS2 proteins in CW and five in EC that bound to FN were observed (Figures 1F,H)
It has been demonstrated that the pathogens can invade host epithelial cells through their own extracellular matrix (ECM)-binding proteins (Molinari et al, 1997; Henderson et al, 2011).The interactions between host ECM proteins and bacterial surface adhesins is a necessary step in the colonization of epithelial surfaces, and has been implicated in bacterial invasion of host cells (Tamura et al, 1994)
Summary
Streptococcus suis (SS) is an important pathogen in swine and human that causes septicemia, meningitis, arthritis, and pneumonia (Gottschalk and Segura, 2000). It is a potential threat of significance to public health, for humans can be infected with the bacteria through skin wounds or the consumption of raw pork (Kay et al, 1995; Yu et al, 2005; Gottschalk et al, 2007). The ECM-binding proteins of bacteria usually make great contributions to the infection and host-pathogen interactions. Large scale identification of the LN- and FNbinding proteins of SS2 which interact with host cells may provide a global view of the pathogenesis of SS2-induced infection
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