Full-Length Galectin-3 Is Required for High Affinity Microbial Interactions and Antimicrobial Activity

While adaptive immunity enables the recognition of a wide range of microbial antigens, immunological tolerance limits reactively toward self to reduce autoimmunity. Some bacteria decorate themselves with self-like antigens as a form of molecular mimicry to limit recognition by adaptive immunity. Recent studies suggest that galectin-4 (Gal-4) and galectin-8 (Gal-8) may provide a unique form of innate immunity against molecular mimicry by specifically targeting microbes that decorate themselves in self-like antigens. However, the binding specificity and antimicrobial activity of many human galectins remain incompletely explored. In this study, we defined the binding specificity of galectin-3 (Gal-3), the first galectin shown to engage microbial glycans. Gal-3 exhibited high binding toward mammalian blood group A, B, and αGal antigens in a glycan microarray format. In the absence of the N-terminal domain, the C-terminal domain of Gal-3 (Gal-3C) alone exhibited a similar overall binding pattern, but failed to display the same level of binding for glycans over a range of concentrations. Similar to the recognition of mammalian glycans, Gal-3 and Gal-3C also specifically engaged distinct microbial glycans isolated and printed in a microarray format, with Gal-3 exhibiting higher binding at lower concentrations toward microbial glycans than Gal-3C. Importantly, Gal-3 and Gal-3C interactions on the microbial microarray accurately predicted actual interactions toward intact microbes, with Gal-3 and Gal-3C displaying carbohydrate-dependent binding toward distinct strains of Providentia alcalifaciens and Klebsiella pneumoniae that express mammalian-like antigens, while failing to recognize similar strains that express unrelated antigens. While both Gal-3 and Gal-3C recognized specific strains of P. alcalifaciens and K. pneumoniae, only Gal-3 was able to exhibit antimicrobial activity even when evaluated at higher concentrations. These results demonstrate that while Gal-3 and Gal-3C specifically engage distinct mammalian and microbial glycans, Gal-3C alone does not possess antimicrobial activity.

Carbohydrate-binding specificity of concanavalin lectin from Canavalia spp.

The carbohydrate-binding specificity of lectins from the four accessions of Canavalia seeds (C. ensiformis (ConA), C. cathartica (ConC), C. gladiata (ConG) and C. rosea (ConM) was studied by hemagglutination inhibition assay using monosaccharides, disaccharides and sugar derivatives. Canavalia seed extracts subjected to ammonium sulphate precipitation and the fraction with higher specific activity were analysed for carbohydrate-binding specificity and its mitogenic potential. All four lectins exhibited similar carbohydrate-binding specificity in agglutination inhibition which is in line with docking experiments. Dmannose and D-maltose were highly specific than other sugars. The results of computational method revealed differences in the affinity towards various carbohydrates. Mitogenic activity of all four lectins in human lymphocytes showed varied mitotic index. Among the four lectins studied, binding affinity to mannose and proliferation index was in the order ConG>ConA>ConM>ConC accounting to the efficacy of biological functions of highly similar analogues.