title: Necessity of the stalk region for immunoglobulin E interaction with CD23.
abstract: Previously, a soluble mouse CD23 chimera, composed of an N-terminal trimeric isoleucine zipper motif (lz) followed by the entire extracellular region (amino acids 48-331) of CD23 (lz-CD2348-331), was prepared and exhibited strong binding to rodent immunoglobulin E (IgE). In the current study, we report the construction of a similar human chimeric protein (lz-huCD2345-321), as well as a series of murine chimeric lz-CD23 mutants with incremental portions of stalk deleted, to further investigate the role of the stalk region in mediating the CD23-IgE interaction. All chimeric proteins were designed such that the predicted heptad structure of the stalk was retained. IgE binding, as determined by the capacity to inhibit 125I-IgE from binding to FcepsilonRI-bearing RBL-2H3 cells, and by surface plasmon-resonance analysis using an IgE-coated sensor chip, was unchanged from the original lz chimera and the binding parameters were similar to those of cell-surface CD23. The minimal murine chimera that retained IgE-binding activity was lz-CD23139-331, which still contains 35 amino acids of the stalk region. When the lz motif was linked to CD23 amino acid 157 (or higher), significant IgE-binding capacity was lost. With human lz-CD23, as with mouse, deletion of the stalk greatly reduced IgE-binding ability. In summary, the data support the concept that at least a portion of the stalk region of CD23 plays a crucial role in maintaining high-affinity/avidity interaction with IgE. The lz-CD23 constructs represent a possible alternative for both blocking the IgE/FcepsilonRI interaction and inhibiting IgE production by B lymphocytes.
authors.affiliation: Department of Microbiology and Immunology, Virginia Commonwealth University, Richmond, VA 23298, USA.
authors.forename: Bing-Hung
authors.initials: BH
authors.lastname: Chen
authors.forename: Check
authors.initials: C
authors.lastname: Ma
authors.forename: Timothy H
authors.initials: TH
authors.lastname: Caven
authors.forename: Yee
authors.initials: Y
authors.lastname: Chan-Li
authors.forename: Andrew
authors.initials: A
authors.lastname: Beavil
authors.forename: Rebecca
authors.initials: R
authors.lastname: Beavil
authors.forename: Hannah
authors.initials: H
authors.lastname: Gould
authors.forename: Daniel H
authors.initials: DH
authors.lastname: Conrad
mesh_terms: D000818:Animals; D000915:Antibody Affinity; D001667:Binding, Competitive; D006801:Humans; D007073:Immunoglobulin E / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D051381:Rats; D017455:Receptors, IgE / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013329:Structure-Activity Relationship; D020349:Surface Plasmon Resonance; D014407:Tumor Cells, Cultured
chemical_list: D017455:Receptors, IgE; D011993:Recombinant Fusion Proteins; D007073:Immunoglobulin E