Understanding the activation and microbial-binding properties of x-type lectin family members
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Intelectins are a novel class of X-type lectins with roles in innate immunity. The first X-type lectin was identified from Xenopus laevis oocytes and embryos. This X. laevis cortical granule lectin (XCGL-1 or XL-35) crosslinks the jelly-coat protein surrounding the oocyte to prevent polyspermy. Following the discovery of XCGL-1, two human intelectin (hIntL-1 and -2) members were identified. The goal of this dissertation was two-fold. First, we investigated the signaling mechanisms for IL-13 induced hIntL-1 activation using a colon cancer cell line. Here, IL-13 activated both the MAPK and JAK/STAT pathways to induce the expression and secretion of hIntL-1. Pharmacologically inhibiting either the MAPK or JAK/STAT pathway prevented IL-13-induced secretion of hIntL-1, suggesting hIntL-1 activation is co-dependent on both pathways. Second, we sought to determine the glycan-binding specificities of members of the X-type lectin family. Using glycan microarrays, we observed specific interactions between recombinant hIntL-1 and -2 and pathogenic microbial glycans isolated from Proteus vulgaris, Streptococcus pneumoniae, Proteus mirabilis, and Klebsiella pneumoniae. Whereas hIntL-1 showed binding to microbial glycans containing β-galactofuranose (β-Galf) and glycerol phosphate (GroP) moieties, hIntL-2 showed minimal binding to glycans with β-Galf and only in specific glycans did it bind GroP. XCGL-1 interacted with the majority of the S. pneumoniae glycans on the array, but not with the affinity of hIntL-1 or -2. Molecular dynamics were employed to validate the interactions between Galf and hIntL-1 and -2. A simulation of β-Galf and hIntL-1 indicated a stable interaction with β-Galf remaining bound to hIntL-1. In contrast, hIntL-2 was unable to bind β-Galf for the duration of the simulation. Amino acid sequence alignment of the three lectins showed multiple key ligand-binding residue substitutions, suggesting these residues are critical for glycan specificity and affinity. Together, these findings examine the unique activation and differential glycan-binding properties of intelectins, and further support their role in the immune system as microbial recognition molecules.