Abstract
BackgroundThe M-like protein, also known as SzP, is expressed on the surface of Streptococcus equi subsp. zooepidemicus (S. zooepidemicus). Previous studies demonstrated that SzP is similar to M protein of group A Streptococcus in the structure and characteristics of antiphagocytosis. The M protein is an adhesin that can bind to the host cells, however it is not known whether the SzP of S. zooepidemicus also functions as an adhesin. We conducted an investigation to determine SzP as an adhesin, and one SzP epitope was identified to be responsible for mediating binding to HEp-2 cells.MethodsThe gene encoding SzP was expressed in E. coli, and the purified recombinant SzP (rSzP) was recognized by rabbit anti-S. zooepidemicus antibodies using immunoblot. Furthermore, the adherence of S. zooepidemicus to HEp-2 cells was inhibited by anti-rSzP antibodies in a dose-dependent manner. We employed a random 12-peptide phage display library for screening of immunodominant mimics of the SzP, which were recognized by an anti-SzP specific monoclonal antibody (mAb 2C8). Initial positive phage clones were identified by ELISA, followed by assays to determine the adherence-inhibiting ability of the peptide.ResultsTen out of fourteen selected positive clones showed high reactivity that effectively inhibited the binding of mAb 2C8 to rSzP. The motif XSLSRX was highly conserved among six of the ten clones.ConclusionCollectively, our findings suggest that the motif XSLSRX may represent an immunodominant mimic epitope of the SzP of S. zooepidemicus strain ATCC 35246, and that the same epitope may be used to mediate SzP binding to HEp-2 cells.
Highlights
The M-like protein, known as SzP, is expressed on the surface of Streptococcus equi subsp. zooepidemicus (S. zooepidemicus)
Out of the 12 monoclonal antibodies,only 2C8 was able to Molecular characterization of M-like protein Nucleotide sequence analysis revealed a single open reading frame in the szp gene of S. zooepidemicus ATCC 35246 isolated from pigs; translation of this open reading frame revealed a protein of 379 amino acids
The szp gene showed 86.9% homology at the nucleotide level with the szp gene of S. zooepidemicus W60 strain isolated from the horse, and 29.4% homology with the M protein gene of SFDigSu-PreAG1E analysis and western blot of recombinant SzP (rSzP) SDS-PAGE analysis and western blot of rSzP
Summary
The M-like protein, known as SzP, is expressed on the surface of Streptococcus equi subsp. zooepidemicus (S. zooepidemicus). The M-like protein, known as SzP, is expressed on the surface of Streptococcus equi subsp. Previous studies demonstrated that SzP is similar to M protein of group A Streptococcus in the structure and characteristics of antiphagocytosis. The M protein is an adhesin that can bind to the host cells, it is not known whether the SzP of S. zooepidemicus functions as an adhesin. Zooepidemicus (S. zooepidemicus), which belongs to Lancefield group C streptococci, is an important animal pathogen, especially in horse [1]. It has a broad host spectrum and occasionally infects humans. In China, S. zooepidemicus is the main pig pathogen. In the summer of 1975, a S. zooepidemicus disease outbreak occurred among pigs in the Sichuan province, China. Clinical symptoms of the diseased pigs included painful swelling of the joints, respiratory distur-
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