UMMS Affiliation

Department of Medicine, Division of Infectious Diseases and Immunology

Publication Date


Document Type



Biochemistry | Cell Biology | Immunology and Infectious Disease | Molecular Biology


Lipopolysaccharide (LPS) activates innate immune responses through TLR4.MD-2. LPS binds to the MD-2 hydrophobic pocket and bridges the dimerization of two TLR4.MD-2 complexes to activate intracellular signaling. However, exactly how lipid A, the endotoxic moiety of LPS, activates myeloid lineage cells remains unknown. Lipid IV(A), a tetra-acylated lipid A precursor, has been used widely as a model for lipid A activation. For unknown reasons, lipid IV(A) activates proinflammatory responses in rodent cells but inhibits the activity of LPS in human cells. Using stable TLR4-expressing cell lines and purified monomeric MD-2, as well as MD-2-deficient bone marrow-derived macrophages, we found that both mouse TLR4 and mouse MD-2 are required for lipid IV(A) activation. Computational studies suggested that unique ionic interactions exist between lipid IV(A) and TLR4 at the dimerization interface in the mouse complex only. The negatively charged 4'-phosphate on lipid IV(A) interacts with two positively charged residues on the opposing mouse, but not human, TLR4 (Lys(367) and Arg(434)) at the dimerization interface. When replaced with their negatively charged human counterparts Glu(369) and Gln(436), mouse TLR4 was no longer responsive to lipid IV(A). In contrast, human TLR4 gained lipid IV(A) responsiveness when ionic interactions were enabled by charge reversal at the dimerization interface, defining the basis of lipid IV(A) species specificity. Thus, using lipid IV(A) as a selective lipid A agonist, we successfully decoupled and coupled two sequential events required for intracellular signaling: receptor engagement and dimerization, underscoring the functional role of ionic interactions in receptor activation.


Lipopolysaccharide, innate immune responses, Lipid A, intracellular signaling

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© 2010 by The American Society for Biochemistry and Molecular Biology, Inc. Publisher PDF posted after 12 months as allowed by the publisher's author rights policy at

DOI of Published Version



This research was originally published in: J Biol Chem. 2010 Mar 19;285(12):8695-702. doi: 10.1074/jbc.M109.075127. Epub 2009 Dec 15. Link to article on publisher's site

Journal/Book/Conference Title

The Journal of biological chemistry

Related Resources

Link to Article in PubMed

PubMed ID