Electrostatic contributions drive the interaction between Staphylococcus aureus protein Efb‐C and its complement target C3d

The C3–inhibitory domain of Staphylococcus aureus extracellular fibrinogen‐binding protein (Efb‐C) defines a novel three‐helix bundle motif that regulates complement activation. Previous crystallographic studies of Efb‐C bound to its cognate subdomain of human C3 (C3d) identified Arg‐131 and Asn‐138 of Efb‐C as key residues for its activity. In order to characterize more completely the physical and chemical driving forces behind this important interaction, we employed in this study a combination of structural, biophysical, and computational methods to analyze the interaction of C3d with Efb‐C and the single‐point mutants R131A and N138A. Our results show that while these mutations do not drastically affect the structure of the Efb‐C/C3d recognition complex, they have significant adverse effects on both the thermodynamic and kinetic profiles of the resulting complexes. We also characterized other key interactions along the Efb‐C/C3d binding interface and found an intricate network of salt bridges and hydrogen bonds that anchor Efb‐C to C3d, resulting in its potent complement inhibitory properties.