Interaction of the LPS-Binding Protein with Host Cell Membranes: Specificity and Biological Implications

Objectives: Lipopolysaccharide-binding protein (LBP) plays a key role in the innate immune response to bacterial infections. Based on LBP-mediated transport of lipopolysaccharide (LPS) to the cellular LPS-receptor TLR4/MD-2, LBP sensitizes immune cell responses to LPS. Besides binding to its ligand LPS, LBP has been demonstrated to interact also with phospholipids. In this study we have addressed the interaction of LBP with the membrane of host cells.Methods: Employing model membranes resembling the eukaryotic cytoplasmic membrane we characterized LBP membrane interaction by flow cytometry, FRET-spectroscopy, IR-spectroscopy, SAXS, and confocal microscopy. LBP binding, uptake and cellular localization was investigated in primary human monocytes, macrophages and in HEK293 cell lines.Results: The investigation of DOPC:SM:CHOL liposome revealed LBP membrane partitioning, its consequences on membrane structure (SAXS at beamline P12, DESY) and biophysical behaviour. Analysis of mononuclear cells from healthy donors showed the presence of endogenous LBP on the cell membrane of monocytes. Association of LBP with distinct membrane domains is shown in analysis of DRM membrane fractions from primary monocytes. Studies on HEK293 cells and human macrophages demonstrate that LBP-binding to host cell membranes occurs independently of the TLR4-receptor and is involved in the uptake and intracellular transport of LPS.Conclusions: In addition to its function in serum, LBP is also abundant in the cytoplasmic membrane and in intracellular compartments of monocytes and macrophages. Cell interaction, localization and intracellular LPS transport by LBP occur independent of TLR4, suggesting a new role of LBP in intracellular LPS-trafficking and signalling.This work was funded in parts by Leibniz-Graduate School “Model systems for infectious diseases” and the Cluster of Excellence “Inflammation at Interfaces” project EXC306OTP4 to A.B.S.