Abstract
Human leukocyte immunoglobulin-like receptors (LILRs) typically regulate immune activation by binding to the human leukocyte antigen class I molecules. LILRA2, a member of the LILR family, was recently reported to bind to other unique ligands, the bacterially degraded Igs (N-truncated Igs), for the activation of immune cells. Therefore, LILRA2 is currently attracting significant attention as a novel innate immune receptor. However, the detailed recognition mechanisms required for this interaction remain unclear. In this study, using several biophysical techniques, we uncovered the molecular mechanism of N-truncated Ig recognition by LILRA2. Surface plasmon resonance analysis disclosed that LILRA2 specifically binds to N-truncated Ig with weak affinity (K(d) = 4.8 μm) and fast kinetics. However, immobilized LILRA2 exhibited a significantly enhanced interaction with N-truncated Ig due to avidity effects. This suggests that cell surface-bound LILRA2 rapidly monitors and identifies bi- or multivalent abnormal N-truncated Igs through specific cross-linking to induce immune activation. Van't Hoff analysis revealed that this interaction is enthalpy-driven, with a small entropy loss, and results from differential scanning calorimetry indicated the instability of the putative LILRA2-binding site, the Fab region of the N-truncated Ig. Atomic force microscopy revealed that N truncation does not cause significant structural changes in Ig. Furthermore, mutagenesis analysis identified the hydrophobic region of LILRA2 domain 2 as the N-truncated Ig-binding site, representing a novel ligand-binding site for the LILR family. These results provide detailed insights into the molecular regulation of LILR-mediated immune responses targeting ligands that have been modified by bacteria.