A novel system for convenient detection of low-affinity receptor-ligand interactions: Chelator-lipid lipsomes engrafted with recombinant CD4 bind to cells expressing MHC class II
Date
2001
Authors
Van Broekhoven, Christina
Altin, Joseph
Journal Title
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Volume Title
Publisher
Blackwell Publishing Ltd
Abstract
The interactions of cell surface receptors with their ligands, crucial for initiating many immunological responses, are often stabilized by receptor dimerization/oligomerization, and by multimeric interactions between receptors on one cell with their ligands or cognate receptors on the apposing cell. Current techniques for studying receptor-ligand interactions, however, do not always allow receptors to move laterally to enable dimerization/oligomerization, or to interact multimerically with ligands on cell surfaces. For these reasons detection of low-affinity receptor-ligand interactions has been difficult. Utilizing a novel chelator-lipid, nitrilotriacetic acid di-tetradecylamine (NTA-DTDA), we have developed a convenient liposome system for directly detecting low-affinity receptor-ligand interactions. Our studies using recombinant soluble forms of murine CD40 and B7.1, and murine and human CD4, each possessing a hexhistidine tag, showed that these proteins can be anchored or 'engrafted' directly onto fluorescently labelled liposomes via a metal-chelating linkage with NTA-DTDA, permitting them to undergo dimerization/oligomerization and multimeric binding with ligands on cells. Fluorescence-activated cell sorter (FACS) analyses demonstrated that while there is little if any binding of soluble forms of murine CD40 and B7.1, and murine and human CD4 to cells, engrafted liposomes bind specifically to cells expressing the appropriate cognate receptor, often giving a fluorescence 4-6-fold above control cells. Such liposomes could detect directly the low-affinity interaction of murine CD40 and B7.1 with CD154- and CD28-expressing cells, respectively, and the interaction of CD4 with MHC Class II, which has hitherto defied direct detection except through mutational analysis and mAb blocking studies.
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Keywords
Keywords: B7 antigen; CD28 antigen; CD4 antigen; CD40 antigen; cell surface receptor; chelating agent; liposome; major histocompatibility antigen class 2; nitrilotriacetic acid; recombinant protein; animal cell; article; binding affinity; controlled study; dimeriza CD4-MHC class II; Cell surface receptors; Chelator-lipid liposomes; Low-affinity interactions
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Source
Immunology and Cell Biology
Type
Journal article
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2037-12-31