Smallpox inhibitor of complement enzymes (SPICE): regulation of complement activation on cells and mechanism of its cellular attachment.
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Despite eradication of smallpox three decades ago, public health concerns remain due to its potential use as a bioterrorist weapon. Smallpox and other orthopoxviruses express virulence factors that inhibit the host's complement system. In this study, our goals were to characterize the ability of the smallpox inhibitor of complement enzymes, SPICE, to regulate human complement on the cell surface. We demonstrate that SPICE binds to a variety of cell types and that the heparan sulfate and chondroitin sulfate glycosaminoglycans serve as attachment sites. A transmembrane-engineered version as well as soluble recombinant SPICE inhibited complement activation at the C3 convertase step with equal or greater efficiency than that of the related host regulators. Moreover, SPICE attached to glycosaminoglycans was more efficient than transmembrane SPICE. We also demonstrate that this virulence activity of SPICE on cells could be blocked by a mAb to SPICE. These results provide insights related to the complement inhibitory activities of poxviral inhibitors of complement and describe a mAb with therapeutic potential. (+info)
Functional basis of protection against age-related macular degeneration conferred by a common polymorphism in complement factor B.
(10/299)
We have investigated the development of substrate-based inhibitors of complement enzymes. Sequences around the scissile Arg77-Ser78 bond of C3 have been synthesized and tested as inhibitors of C3 convertase. The best inhibition was found with the tetrapeptide Ac-Arg-Ser-Asn-Leu-OH (H-576); extending this sequence in either direction reduced inhibitory activity. Preliminary experiments with peptides in which the scissile bond--CO--NH--was replaced with non-hydrolysable moieties such as--CO--CH2--(H-497) and--CH2--NH--(H-336) failed to show enhanced inhibition. One of the longer chain inhibitors H-416 containing DArg77-Ser78 was unexpectedly found to potentiate iC3 cleavage by factors I and H but did not inhibit the intact alternative pathway. The same peptide also bound to factor H. It is concluded that the binding requirements of the C3 convertase are more sophisticated than can be satisfactorily imitated simply by linear sequences around the scissile bond of C3. (+info)
Inhibition of the alternative C3 convertase and classical C5 convertase of complement by group A streptococcal M protein.
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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. (+info)
Disease-causing mutations in genes of the complement system.
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