Analysis of the Role of N-Linked Glycans on Human CD1d Molecules

CD1d is a cell-surface expressed glycoprotein that presents a variety of lipid and glycolipid antigens to invariant natural killer T-cells (iNKT). We have examined the role of the N-linked glycans on the human CD1d (hCD1d) in modulating the responses of iNKT cells using a combination of cell-free, cell-based assays and in silico analyses. Deficiency of one or more glycans on hCD1d diminished the activation of iNKT cells. Soluble glycan mutants did not have significant differences in terms of binding to the nickel-coated 96-well plates. Our cell-free assay, together with docking and MM-GBSA analyses, confirms that Glycan 1 and Glycan 2 are critical determinants of optimal iNKT cell activation. Taken together, our data emphasized an essential role of the hCD1d N-linked glycans in modulating iNKT cell response.