title: Lipopolysaccharide (LPS)-binding protein accelerates the binding of LPS to CD14.
abstract: CD14 is a 55-kD protein found as a glycosylphosphatidylinositol (GPI)-anchored protein on the surface of monocytes, macrophages, and polymorphonuclear leukocytes, and as a soluble protein in the blood. Both forms of CD14 participate in the serum-dependent responses of cells to bacterial lipopolysaccharide (LPS). While CD14 has been described as a receptor for complexes of LPS with LPS-binding protein (LBP), there has been no direct evidence showing whether a ternary complex of LPS, LBP, and CD14 is formed, or whether CD14 binds LPS directly. Using nondenaturing polyacrylamide gel electrophoresis (native PAGE), we show that recombinant soluble CD14 (rsCD14) binds LPS in the absence of LBP or other proteins. Binding of LPS to CD14 is stable and of low stoichiometry (one or two molecules of LPS per rsCD14). Recombinant LBP (rLBP) does not form detectable ternary complexes with rsCD14 and LPS, but it does accelerate the binding of LPS to rsCD14. rLBP facilitates the interaction of LPS with rsCD14 at substoichiometric concentrations, suggesting that LBP functions catalytically, as a lipid transfer protein. Complexes of LPS and rsCD14 formed in the absence of LBP or other serum proteins strongly stimulate integrin function on PMN and expression of E-selectin on endothelial cells, demonstrating that LBP is not necessary for CD14-dependent stimulation of cells. These results suggest that CD14 acts as a soluble and cell surface receptor for LPS, and that LBP may function primarily to accelerate the binding of LPS to CD14.
authors.affiliation: Laboratory of Cellular Physiology and Immunology, Rockefeller University, New York 10021.
authors.forename: E
authors.initials: E
authors.lastname: Hailman
authors.forename: H S
authors.initials: HS
authors.lastname: Lichenstein
authors.forename: M M
authors.initials: MM
authors.lastname: Wurfel
authors.forename: D S
authors.initials: DS
authors.lastname: Miller
authors.forename: D A
authors.initials: DA
authors.lastname: Johnson
authors.forename: M
authors.initials: M
authors.lastname: Kelley
authors.forename: L A
authors.initials: LA
authors.lastname: Busse
authors.forename: M M
authors.initials: MM
authors.lastname: Zukowski
authors.forename: S D
authors.initials: SD
authors.lastname: Wright
mesh_terms: D000209:Acute-Phase Proteins; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism*; D015214:Antigens, Differentiation, Myelomonocytic / Q000378:metabolism*; D001483:Base Sequence; D016466:CHO Cells; D002352:Carrier Proteins / Q000502:physiology*; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002478:Cells, Cultured; D006224:Cricetinae; D017931:DNA Primers; D019040:E-Selectin; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides / Q000378:metabolism* / Q000494:pharmacology; D008562:Membrane Glycoproteins; D008969:Molecular Sequence Data; D009504:Neutrophils / Q000276:immunology; D009895:Opsonin Proteins; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D000099231:Lipopolysaccharide-Binding Protein
chemical_list: D000209:Acute-Phase Proteins; D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D002352:Carrier Proteins; D015815:Cell Adhesion Molecules; D017931:DNA Primers; D019040:E-Selectin; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D009895:Opsonin Proteins; D011994:Recombinant Proteins; D000099231:Lipopolysaccharide-Binding Protein