Tianjin University of Science and Technology, Tianjin, P.R. China.
J Insect Sci. 2010;10:79. doi: 10.1673/031.010.7901.
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 degrees C and in pH ranging from 4 to 8. Salts including FeCl(3) and MnCl(2) inhibited the haemagglutinating process, whereas NaCl, KCl, CaCl(2), MgCl(2), ZnCl(2), and AlCl(3) 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 beta-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 IC(50) of 5.7 and 6.7 at 24 h, 5.5 and 6.4 at 36 h, 5.2 and 6.5 microM at 48 h, respectively.
在本工作中，从家蝇（双翅目：蝇科）蛹中纯化了一种具有新型 N 末端序列的 D-半乳糖特异性凝集素。通过亲和层析、超滤和 HPLC 进行纯化。家蝇凝集素的血凝活性被 D-半乳糖特异性抑制。该凝集素的血凝活性在高达 65°C 的温度和 pH 值为 4 至 8 的范围内稳定。包括 FeCl3 和 MnCl2 在内的盐抑制了血凝过程，而 NaCl、KCl、CaCl2、MgCl2、ZnCl2 和 AlCl3 则没有。通过 SDS-PAGE，纯化的家蝇蛹凝集素在有无β-巯基乙醇还原的情况下产生一条分子量为 40 kDa 的单带，并且可以用 Alcian Blue 8 GX 染色。通过原子力显微镜观察到纯化的凝集素的形态，表明家蝇凝集素是一种 8.27 nm 高的球形糖蛋白，带有 1.41 nm 高的多糖链。此外，用噻唑蓝比色法测定了该凝集素对肿瘤细胞 K562 和 MCF-7 的抗增殖活性，结果表明，抗增殖过程是时间和剂量依赖性的，在 24 h 时的 IC50 分别为 5.7 和 6.7 μM，在 36 h 时分别为 5.5 和 6.4 μM，在 48 h 时分别为 5.2 和 6.5 μM。
