Structural insights into recognition of chemokine receptors by Staphylococcus aureus leukotoxins.

Staphylococcus aureus (SA) leukocidin ED (LukED) belongs to a family of bicomponent pore forming toxins that play important roles in SA immune evasion and nutrient acquisition. LukED targets specific G protein-coupled chemokine receptors to lyse human erythrocytes (red blood cells) and leukocytes (white blood cells). The first recognition step of receptors is critical for specific cell targeting and lysis. The structural and molecular bases for this mechanism are not well understood but could constitute essential information to guide antibiotic development. Here, we characterized the interaction of LukE with chemokine receptors ACKR1, CCR2, and CCR5 using a combination of structural, pharmacological, and computational approaches. First, crystal structures of LukE in complex with a small molecule mimicking sulfotyrosine side chain (p-cresyl sulfate) and with peptides containing sulfotyrosines issued from receptor sequences revealed the location of receptor sulfotyrosine binding sites in the toxins. Then, by combining previous and novel experimental data with protein docking, classical and accelerated weight histogram (AWH) molecular dynamics we propose models of the ACKR1-LukE and CCR5-LukE complexes. This work provides novel insights into chemokine receptor recognition by leukotoxins and suggests that the conserved sulfotyrosine binding pocket could be a target of choice for future drug development.

Molecular insights into mechanisms of GPCR hijacking by Staphylococcus aureus.

Atypical chemokine receptor 1 (ACKR1) is a G protein-coupled receptor (GPCR) targeted by Staphylococcus aureus bicomponent pore-forming leukotoxins to promote bacterial growth and immune evasion. Here, we have developed an integrative molecular pharmacology and structural biology approach in order to characterize the effect of leukotoxins HlgA and HlgB on ACKR1 structure and function. Interestingly, using cell-based assays and native mass spectrometry, we found that both components HlgA and HlgB compete with endogenous chemokines through a direct binding with the extracellular domain of ACKR1. Unexpectedly, hydrogen/deuterium exchange mass spectrometry analysis revealed that toxin binding allosterically modulates the intracellular G protein-binding domain of the receptor, resulting in dissociation and/or changes in the architecture of ACKR1-Gαi1 protein complexes observed in living cells. Altogether, our study brings important molecular insights into the initial steps of leukotoxins targeting a host GPCR.