Ecotin, a microbial inhibitor of serine proteases, blocks multiple complement dependent and independent microbicidal activities of human serum

English version

Autoři: Zoltán Attila Nagy ^aff001; Dávid Szakács ^aff001; Eszter Boros ^aff001; Dávid Héja ^aff001; Eszter Vígh ^aff001; Noémi Sándor ^aff003; Mihály Józsi ^aff003; Gábor Oroszlán ^aff004; József Dobó ^aff004; Péter Gál ^aff004; Gábor Pál ^aff001
Působiště autorů: Department of Biochemistry, ELTE, Eötvös Loránd University, Budapest, Hungary ^aff001; Department of Nephrology, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America ^aff002; Department of Immunology, ELTE, Eötvös Loránd University, Budapest, Hungary ^aff003; Institute of Enzymology, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary ^aff004
Vyšlo v časopise: Ecotin, a microbial inhibitor of serine proteases, blocks multiple complement dependent and independent microbicidal activities of human serum. PLoS Pathog 15(12): e32767. doi:10.1371/journal.ppat.1008232
Kategorie: Research Article
doi: https://doi.org/10.1371/journal.ppat.1008232

Souhrn

Ecotin is a serine protease inhibitor produced by hundreds of microbial species, including pathogens. Here we show, that ecotin orthologs from Escherichia coli, Yersinia pestis, Pseudomonas aeruginosa and Leishmania major are potent inhibitors of MASP-1 and MASP-2, the two key activator proteases of the complement lectin pathway. Factor D is the key activator protease of another complement activation route, the alternative pathway. We show that ecotin inhibits MASP-3, which is the sole factor D activator in resting human blood. In pathway-specific ELISA tests, we found that all ecotin orthologs are potent lectin pathway inhibitors, and at high concentration, they block the alternative pathway as well. In flow cytometry experiments, we compared the extent of complement-mediated opsonization and lysis of wild-type and ecotin-knockout variants of two E. coli strains carrying different surface lipopolysaccharides. We show, that endogenous ecotin provides significant protections against these microbicidal activities for both bacteria. By using pathway specific complement inhibitors, we detected classical-, lectin- and alternative pathway-driven complement attack from normal serum, with the relative contributions of the activation routes depending on the lipopolysaccharide type. Moreover, in cell proliferation experiments we observed an additional, complement-unrelated antimicrobial activity exerted by heat-inactivated serum. While ecotin-knockout cells are highly vulnerable to these activities, endogenous ecotin of wild-type bacteria provides complete protection against the lectin pathway-related and the complement-unrelated attack, and partial protection against the alternative pathway-related damage. In all, ecotin emerges as a potent, versatile self-defense tool that blocks multiple antimicrobial activities of the serum. These findings suggest that ecotin might be a relevant antimicrobial drug target.

Klíčová slova:

Bacterial pathogens – Complement system – Enzyme inhibitors – Flow cytometry – Proteases – Pseudomonas aeruginosa – Complement inhibitors – Yersinia pestis

Zdroje

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