Structural Basis for Ligand Recognition and Discrimination of a Quorum-quenching Antibody

Abstract

In the postantibiotic era, available treatment options for severe bacterial infections caused by methicillin-resistant Staphylococcus aureus have become limited. Therefore, new and innovative approaches are needed to combat such lifethreatening infections. Virulence factor expression in S. aureus is regulated in a cell density-dependent manner using "quorum sensing," which involves generation and secretion of autoinducing peptides (AIPs) into the surrounding environment to activate a bacterial sensor kinase at a particular threshold concentration. Mouse monoclonal antibody AP4-24H11 was shown previously to blunt quorum sensing-mediated changes in gene expression in vitro and protect mice from a lethal dose of S. aureus by sequestering the AIP signal. We have elucidated the crystal structure of the AP4-24H11 Fab in complex with AIP-4 at 2.5 Å resolution to determine its mechanism of ligand recognition. A key Glu H95 provides much of the binding specificity through formation of hydrogen bonds with each of the four amide nitrogens in the AIP-4 macrocyclic ring. Importantly, these structural data give clues as to the interactions between the cognate staphylococcal AIP receptors AgrC and the AIPs, as AP4-24H11⅐AIP-4 binding recapitulates features that have been proposed for AgrC-AIP recognition. Additionally, these structural insights may enable the engineering of AIP cross-reactive antibodies or quorum quenching vaccines for use in active or passive immunotherapy for prevention or treatment of S. aureus infections.

Coagulase-Negative Staphylococcal Strain Prevents Staphylococcus aureus Colonization and Skin Infection by Blocking Quorum Sensing

“…Staphylococcus epidermidis has already been tested in depth (Otto et al, 2001; Otto et al, 1999), and thus we focused on other staphylococcal species. For reporters, we used S. aureus strains representing all four agr types containing P3 promoter YFP-fusions that have been previously used to assess quorum-sensing activity (Kirchdoerfer et al, 2011; Quave et al, 2015). In our assays, Staphylococcus capitis and multiple clinical isolates of Staphylococcus lugdunensis did not affect S. aureus agr activation, while agr P3 inhibition was observed in spent media from Staphylococcus intermedius and Staphylococcus caprae (Fig.…”

Characterization of structural elements in native autoinducing peptides and non-native analogues that permit the differential modulation of AgrC-type quorum sensing receptors in Staphylococcus aureus

“…For AgrC inhibition, we used S. aureus strains (groups I–IV) possessing P3- gfp reporter plasmids (see Experimental Section). 24 In these strains, activation of AgrC by the AIP signal results in AgrA phosphorylation, which in turn binds the P3 promoter and induces gfp transcription (Figure 1). These strains are capable of native AIP production, and thus GFP is produced at quorate cell densities.…”

Structure–Function Analyses of a Staphylococcus epidermidis Autoinducing Peptide Reveals Motifs Critical for AgrC-type Receptor Modulation

“…The S. epidermidis wild-type strain RP62A (Group-I) was cultured in TSB (from Sigma or EMD Millipore). The S. aureus fluorescence reporter strains AH1677 (Group-I), AH430 (Group-II), AH1747 (Group-III), and AH1872 (Group-IV) were cultured in Brain Heart Infusion (BHI, from Teknova) augmented with 10 μg mL −1 of chloramphenicol (65). All cultures were grown at 37 °C with shaking (200 rpm) unless noted otherwise.…”