DC‐SIGN Binds 2‐Fucosyl‐Lactose (2FL) at Concentrations Available in Human Milk

Human epithelial and immune cells express many glycan‐binding proteins (GBPs) and their interactions with glycans in human milk could contribute to immune regulation and infant health. To explore this, DC‐SIGN, which is expressed by dendritic cells and macrophages, was analyzed for glycan binding on a glycan microarray comprised of 247 glycan targets purified from human milk (HMG260 array). We also developed a smaller array with 18 defined glycans (defined array), immobilized in reduced and non‐reduced forms (pyranose form). DC‐SIGN (concentration range of 0.1‐5 µg/ml) bound to a large number of glycans on the HMG260 array; several strong binding glycans were structurally analyzed using metadata‐assisted glycan sequencing (MAGS). MAGS revealed that these DC‐SIGN‐binding glycans were fucosylated, including those of the Lewis blood group. On the defined array, DC‐SIGN strongly bound to 2FL (Fucα1‐2Galβ1‐4Glc) and 3FL (pyranose form) (Galβ1‐4(Fucα1‐3)Glc p ), and weakly bound to LNFPI (Fucα1‐2Galβ1‐3GlcNAcβ1‐3Galβ1‐4Glc). DC‐SIGN bound to the free form of 2FL, as shown by the ability of 2FL to inhibit the binding of DC‐SIGN to immobilized glycans, while lactose was not inhibitory. In addition, polystyrene beads conjugated with 2FL bound DC‐SIGN, as determined by flow cytometry, and such binding was inhibited by free 2FL but not by lactose. These data indicate that DC‐SIGN expressed in infant intestine may be affected by 2FL at concentrations available in milk of breast fed infants. [This work funded in part by NIH Grant GM103694 to RDC and under a research agreement with Abbott Nutrition. DFS and RDC are consultants for Abbott Nutrition.]