Full-length galectin-3 and a galectin-3 N-terminal deletion mutant were produced as previously described (47). Heterologous protein expression was induced in Rosetta Escherichia coli clones by the addition of 0.3 mM IPTG (isopropyl-β-d-thiogalactopyranoside). The proteins were purified from lysates by affinity chromatography using lactosyl sepharose. To eliminate contaminating bacterial endotoxins, the proteins were further purified by polymyxin B affinity chromatography (Sigma-Aldrich). The absence of lipopolysaccharide was confirmed using the ToxinSensor Chromogenic LAL Endotoxin Assay Kit (GenScript). Protein solutions were concentrated by centrifugal filtration (VWR), dialyzed against PBS buffer containing 10% glycerol, and stored at −20°C.

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