Purification and Characterization of a New D-Galactose-Specific Lectin from the Housefly, Musca domestica, and Its Antiproliferative Effect on Human K562 and MCF-7 Tumor Cells

摘要

In the present work, a D-galactose-specific lectin with novel N-terminal sequence was purified from Musca domestica L. (Diptera: Muscidae) pupae. The purification was performed using affinity chromatography, ultra-filtration, and HPLC. The haemagglutinating activity of M. domestica lectin was specifically inhibited by D-galactose. The haemagglutinating activity of this lectin was stable at temperatures up to 65° C and in pH ranging from 4 to 8. Salts including FeCl3 and MnCl2 inhibited the haemagglutinating process, whereas NaCl, KCl, CaCl2, MgCl2, ZnCl2, and AlCl3 did not. By SDS-PAGE, purified M. domestica pupae lectin yielded a single band with a molecular weight of 40 kDa, with or without reduction of β-mercaptoethanol, and it could be stained with Alcian Blue 8 GX. The morphology of purified lectin was observed by atomic force microscopy, which indicated that M. domestica lectin was an 8.27 nm high, globular shaped glycoprotein with a 1.41 nm high polysaccharide chain. In addition, antiproliferative activity of this lectin against tumor cells K562 and MCF-7 was determined with a colorimetric assay using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide, which showed that the antiproliferative process was time- and dose-dependent with an IC50 of 5.7 and 6.7 at 24 h, 5.5 and 6.4 at 36 h, 5.2 and 6.5 µM at 48 h, respectively.