Reviving lost binding sites: Exploring calcium‐binding site transitions between human and murine CD23

Immunoglobulin E (IgE) is a central regulatory and triggering molecule of allergic immune responses. IgE’s interaction with CD23 modulates both IgE production and functional activities.CD23 is a noncanonical immunoglobulin receptor, unrelated to receptors of other antibody isotypes. Human CD23 is a calcium‐dependent (C‐type) lectin‐like domain that has apparently lost its carbohydrate‐binding capability. The calcium‐binding site classically required for carbohydrate binding in C‐type lectins is absent in human CD23 but is present in the murine molecule. To determine whether the absence of this calcium‐binding site affects the structure and function of human CD23, CD23 mutant proteins with increasingly “murine‐like” sequences were generated. Restoration of the calcium‐binding site was confirmed by NMR spectroscopy, and structures of mutant human CD23 proteins were determined by X‐ray crystallography, although no electron density for calcium was observed. This study offers insights into the evolutionary differences between murine and human CD23 and some of the functional differences between CD23 in different species. A second calcium‐binding site was engineered into human CD23, based on the amino acid sequence of murine CD23. Chemical shifts from calcium HSQC NMR titrations verified the presence and activity of the engineered site. Differences between murine and human CD23 suggest that modulation of the interaction between CD23 and IgE by calcium may play a greater role in humans.