Functional recruitment of human complement inhibitor C4B-binding protein to outer membrane protein Rck of Salmonella

PLoS One. 2011;6(11):e27546. doi: 10.1371/journal.pone.0027546. Epub 2011 Nov 10.

Abstract

Resistance to complement mediated killing, or serum resistance, is a common trait of pathogenic bacteria. Rck is a 17 kDa outer membrane protein encoded on the virulence plasmid of Salmonella enterica serovars Typhimurium and Enteritidis. When expressed in either E. coli or S. enterica Typhimurium, Rck confers LPS-independent serum resistance as well as the ability to bind to and invade mammalian cells. Having recently shown that Rck binds the inhibitor of the alternative pathway of complement, factor H (fH), we hypothesized that Rck can also bind the inhibitor of the classical and lectin pathways, C4b-binding protein (C4BP). Using flow cytometry and direct binding assays, we demonstrate that E. coli expressing Rck binds C4BP from heat-inactivated serum and by using the purified protein. No binding was detected in the absence of Rck expression. C4BP bound to Rck is functional, as we observed factor I-mediated cleavage of C4b in cofactor assays. In competition assays, binding of radiolabeled C4BP to Rck was reduced by increasing concentrations of unlabeled protein. No effect was observed by increasing heparin or salt concentrations, suggesting mainly non-ionic interactions. Reduced binding of C4BP mutants lacking complement control protein domains (CCPs) 7 or 8 was observed compared to wt C4BP, suggesting that these CCPs are involved in Rck binding. While these findings are restricted to Rck expression in E. coli, these data suggest that C4BP binding may be an additional mechanism of Rck-mediated complement resistance.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Bacterial Outer Membrane Proteins / immunology
  • Bacterial Outer Membrane Proteins / metabolism*
  • Complement Activation / drug effects
  • Complement C4b / antagonists & inhibitors*
  • Complement C4b / metabolism
  • Complement C4b-Binding Protein / immunology
  • Complement C4b-Binding Protein / metabolism*
  • Complement Inactivator Proteins / immunology
  • Complement Inactivator Proteins / metabolism*
  • Escherichia coli / immunology
  • Escherichia coli / metabolism
  • Flow Cytometry
  • Heparin / pharmacology
  • Humans
  • Protein Binding
  • Salmonella / immunology*
  • Salmonella Infections / immunology*
  • Salmonella Infections / metabolism
  • Sodium Chloride / pharmacology
  • Virulence / drug effects

Substances

  • Bacterial Outer Membrane Proteins
  • Complement C4b-Binding Protein
  • Complement Inactivator Proteins
  • Sodium Chloride
  • Complement C4b
  • Heparin