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Streptococcal inhibitor of complement (SIC) is a 31 kDa extracellular protein produced by a few highly virulent strains of Streptococcus pyogenes (in particular the M1 strain). It has been shown additionally to inhibit four further components of the mucosal innate response—lysozyme, secretory leucocyte proteinase inhibitor, human α-defensin 1 and the cathelicidin LL-37 which are all bactericidal against Group A Streptococci (GAS). We now show that SIC also inhibits variably the antibacterial action of hBD-1, -2 and -3. By enzyme-linked immunosorbent assay (ELISA), SIC binds strongly to hBD-2 and hBD-3, but not at all to hBD-1. Investigation of the antimicrobial action of β-defensins hBD-1, -2 and -3 against GAS in two different buffer systems shows that both the killing efficiencies of all three defensins, and the binding of SIC to them, occurs more efficiently in 10 m m Tris buffer than in 10 m m phosphate. The lower ionic strength of the Tris buffer may underlie this effect. hBD-1 kills the M1 strain of GAS only in 10 m m Tris, but is able to kill an M6 (SIC negative) strain in 10 m m phosphate. The inhibition of hBD-3 by SIC is clearly of physiological relevance, that of hBD-2 is likely to be so, but the inhibition of hBD-1 occurs only at lower ionic strength than is likely to be encountered in vivo. Elastase digestion of SIC yields three major fragments of MW 3·843 kDa comprising residues 1–33 (fragment A); 10·369 kDa comprising residues 34–126 (fragment B); and MW 16·487 kDa, comprising residues 127–273 (fragment C). By ELISA, only fragment B binds to hBD-2 and hBD-3 and this may indicate the inhibitory portion of the SIC molecule. [ABSTRACT FROM AUTHOR]

Summary Streptococcal inhibitor of complement (SIC) was first described in 1996 as a putative inhibitor of the membrane attack complex of complement (MAC). SIC is a 31 000 MW protein secreted in large quantities by the virulent Streptococcus pyogenes strains M1 and M57, and is encoded by a gene which is extremely variable. In order to study further the interactions of SIC with the MAC, we have made a recombinant form of SIC (rSIC) in Escherichia coli and purified native M1 SIC which was used to raise a polyclonal antibody. SIC prevented reactive lysis of guinea pig erythrocytes by the MAC at a stage prior to C5b67 complexes binding to cell membranes, presumably by blocking the transiently expressed membrane insertion site on C7. The ability of SIC and clusterin (another putative fluid phase complement inhibitor) to inhibit complement lysis was compared, and found to be equally efficient. In parallel, by enzyme-linked immunosorbent assay both SIC and rSIC bound strongly to C5b67 and C5b678 complexes and to a lesser extent C5b-9, but only weakly to individual complement components. The implications of these data for virulence of SIC-positive streptococci are discussed, in light of the fact that Gram-positive organisms are already protected against complement lysis by the presence of their peptidoglycan cell walls. We speculate that MAC inhibition may not be the sole function of SIC. [ABSTRACT FROM AUTHOR]