Expression, purification and functional characterization of Galectin-12 from Mus musculus

MALLER, SEBASTIÁN M.; CAGNONI, ALEJANDRO J.; RABINOVICH, GABRIEL A.; DI LELLA, SANTIAGO; MARIÑO, KARINA V.

Guanajuato

Congreso; 3er Congreso Latinoamericano de Gliobiología; 2015

Adipocytes, the primary cellular component of adipose tissue, store lipids and release free fatty acids and other lipid moieties; these specialized cells are also involved in essential processes for energy homeostasis. Inflammation-induced obesity disrupts this balance, contributing to certain metabolic conditions such as insulin resistance, metabolic syndrome and type-2 diabetes. Galectin-12 (Gal-12) is a tandem-repeat member of the galectin family preferentially expressed in lipid droplets of adipocytes, and its ability to supress lipolysis has attracted considerable interest. Understanding the mechanisms behind the anti-lipolytic activity of Gal-12 may be useful for the development of specific therapies for obesity-related disorders. To further evaluate its physiologic function in health and disease, we have expressed and purified recombinant murine Gal-12 (mGal-12) in E. coli and analyzed its glycan-binding preferences and biological activity. Using a solid phase assay specifically developed for this lectin, we analyzed mGal-12 affinity for different glycans including β-lactose, 3'-sialyllactose, 6'-sialyllactose, 3-fucosyllactose, and LewisX (Lex) hexaose. We demonstrated a particular pattern of recognition for this galectin, as it showed clear preference for LeX moieties [Galβ1-4(Fucα1-3)GlcNAcβ1-3] and exhibited no considerable affinity to β-lactose. Since this protein has never been crystallized, we also generated and refined a molecular model by homology to other tandem-repeat type galectins, a valuable tool for studying ligand binding through molecular docking simulations. These biochemical studies lay the groundwork for unraveling the role of Gal-12 in inflammation-induced obesity and metabolic diseases.