Identification and characterization of a lipopolysaccharide binding, anti-inflammatory protein from <i>Parabacteroides distasonis</i>

Gut bacteria are well known to impact a diverse array of acute and chronic diseases. One mechanism by which gut bacteria can impact health is by modulating inflammation. We and others previously found that membrane components of the gram-negative bacterium <i>Parabacteroides distasonis</i> (Pd) can suppress proinflammatory signaling <i>in vitro</i> and reduce intestinal inflammation in mouse models. Here, we sought to identify anti-inflammatory proteins from the membrane fraction of Pd. Using size exclusion chromatography, cell-based assays, fluorescence anisotropy, pull-down assays, and proteomics, we identified a putative anti-inflammatory protein named DUF4925 domain-containing protein (Protein ID: A6LA28, Gene ID: BDI_0772). Recombinant A6LA28 dose-dependently suppressed LPS induced IL-8 production from HT-29 and SW-480 cells, IL-6 and TNFα from splenocytes and blocked LPS binding to HT-29 and HEK reporter cells. Deletion of the C-terminal 201 amino acids, which are predicted to form a LPS binding pocket, destroyed its function, whereas deletion of the N-terminal 133 amino acids had little impact. Although its native function in Pd remains unclear, these data indicate that A6LA28 can bind to LPS, thereby preventing TLR4-mediated cytokine production. Further work is required to determine whether A6LA28 contributes to the anti-inflammatory capacity of Pd <i>in vivo</i> and to evaluate the therapeutic potential of this protein.