Abstract
Streptococcal pyrogenic exotoxin B (SPE B), a cysteine protease, is an important virulence factor in group A streptococcal (GAS) infection. SPE B binds and cleaves antibody isotypes and further impairs the immune system by inhibiting complement activation. In this study, we examined the antibody-binding site of SPE B and used it to block SPE B actions during GAS infection. We constructed different segments of the spe B gene and induced them to express different recombinant fragments of SPE B. Using an enzyme-linked immunosorbent assay (ELISA), we found that residues 345–398 of the C-terminal domain of SPE B (rSPE B345–398), but not the N-terminal domain, was the major binding site for antibody isotypes. Using a competitive ELISA, we also found that rSPE B345–398 bound to the Fc portion of IgG. The in vitro functional assays indicate that rSPE B345–398 not only interfered with cleavage of antibody isotypes but also interfered with SPE B-induced inhibition of complement activation. Immunization of BALB/c mice using rSPE B345–398 was able to induce production of a high titer of anti-rSPE B345–398 antibodies and efficiently protected mice from GAS-induced death. These findings suggest that SPE B uses its C-terminal domain to bind the Fc portion of IgG and that immunization of mice with this binding domain (rSPE B345–398) could protect mice from GAS infection.
Highlights
Streptococcus pyogenes is an important human pathogen that causes a variety of diseases, including pharyngitis, cellulitis, impetigo, scarlet fever, necrotizing fasciitis, puerperal sepsis, and streptococcal toxic shock syndrome (STSS) [1,2,3]
When compared with protein L-purified human IgG, IgM and IgA, we found that fraction 13 (F13) contained human IgG, IgM, and IgA and that the amount of IgG in the F13 fraction is most abundant, IgM is less, and IgA is least abundant (Figure 1A–D)
The results indicate that protein A but not protein L could inhibit IgG binding to rSPE B345–398 (Figure 5A) and that the competitive inhibition was dose-dependent for protein A (Figure 5B)
Summary
Streptococcus pyogenes (group A streptococcus; GAS) is an important human pathogen that causes a variety of diseases, including pharyngitis, cellulitis, impetigo, scarlet fever, necrotizing fasciitis, puerperal sepsis, and streptococcal toxic shock syndrome (STSS) [1,2,3]. Several virulence factors have been reported that contribute to evasion of host immunity by GAS These factors consist of the cell surface M protein, M-like protein, the hyaluronic acid capsule, the streptococcal inhibitor of complement, and C5a peptidase [5,6,7,8,9], as well as secreted exotoxins and enzymes such as streptococcal pyrogenic exotoxin B (SPE B), IdeS (IgG-degrading enzyme of S. pyogenes), endo-b-N-acetylglucosaminidase (EndoS), and DNases [10,11,12,13,14]. Patients with lower antibody levels against SPE B are more likely to succumb to invasive GAS disease [30] Taken together, these reports indicate that SPE B is a critical virulence factor in GAS infection
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