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Inhibition of the alternative C3 convertase and classical C5 convertase of complement by group A streptococcal M
K Hong1, T Kinoshita, J Takeda
1Department of Bacteriology, Osaka University Medical School, Japan.
Abstract:
When Streptococcus pyogenes group A type 3 strain C203 (M+) and its M-protein-lacking derivative, strain C203S (M-), were treated with normal human serum in the presence of magnesium-EGTA [ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid], virulent M+ bacteria bound only 10 to 30% as much C3 and factors B and P as did avirulent M- bacteria. After treatment of M+ bacteria with trypsin, which inactivates M protein, their binding of these substances was similar to that of M- bacteria. Pretreatment of M+ bacteria with the Fab fragment of rabbit immunoglobulin G anti-M antibody also increased their binding of C3 in the absence of Ca2+. Therefore, M protein inhibits the alternative C3 convertase. In contrast, in the presence of Ca2+ and Mg2+, M+ bacteria bound 75% as much C3 as M- bacteria. This binding was mostly mediated by classical pathway activation, because M+ bacteria bound much smaller amounts of factors B and P than did M- bacteria but consumed amounts of C4 and C2 comparable to those consumed by M- bacteria. On the other hand, the amount of C5 bound to M+ bacteria was much less than that bound to M- bacteria, and the consumption of C5 and C8 by M+ bacteria was also much less than that by M- bacteria. Therefore, M protein does not inhibit the classical C3 convertase but does inhibit the classical C5 convertase. When M+ and M- streptococci were incubated with normal human serum containing radiolabeled C3 in the presence of Ca2+ and Mg2+, more than 85% of the C3 bound to either type of streptococcus was extractable by sodium dodecyl sulfate and alkali treatment. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of the C3 extracted from both strains showed that it was mostly C3b and iC3b. The proportions of C3b and iC3b, respectively, were 7.5 and 71.9% on M+ bacteria and 18.9 and 58.4% on M- bacteria. These results support and extend previous findings that the antiphagocytic activity of streptococcal M protein may be due to complement inhibition mediated by the binding of factor H.
Insights
Streptococcus pyogenes M protein inhibits complement pathways, specifically the alternative C3 and classical C5 convertases. This M protein binding of factor H contributes to the antiphagocytic activity of M+ bacteria.
Area of Science:
- Microbiology
- Immunology
- Biochemistry
Background:
- Streptococcus pyogenes utilizes M protein for virulence and antiphagocytic activity.
- The complement system plays a crucial role in innate immunity against bacterial infections.
Purpose of the Study:
- To investigate the mechanism by which M protein influences complement activation pathways.
- To determine if M protein inhibits complement component C3 convertase or C5 convertase.
Main Methods:
- Treatment of M+ and M- Streptococcus pyogenes strains with normal human serum containing magnesium-EGTA or Ca2+ and Mg2+.
- Quantification of bound complement components (C3, factors B, P, C4, C2, C5, C8) using radiolabeled assays.
- Analysis of bound C3 fragments (C3b, iC3b) via SDS-PAGE after sodium dodecyl sulfate and alkali extraction.
Main Results:
- M protein inhibits the alternative C3 convertase by reducing C3, factor B, and factor P binding in the absence of Ca2+.
- M protein does not inhibit the classical C3 convertase but inhibits the classical C5 convertase.
- M+ bacteria bound significantly less C5 and consumed less C5 and C8 compared to M- bacteria.
- Bound C3 on M+ bacteria was predominantly iC3b, suggesting complement inhibition mediated by factor H.
Conclusions:
- Streptococcal M protein acts as a complement inhibitor, specifically targeting the alternative C3 and classical C5 convertases.
- M protein's antiphagocytic function is likely mediated by its interaction with factor H, leading to complement inhibition.