重组金针菇免疫调节蛋白的纯化及生物活性研究

目的研究重组的金针菇免疫调节蛋白(re FIP-fve)对小鼠脾淋巴细胞的增殖作用、诱导小鼠脾淋巴细胞对白细胞介素-2(IL-2)的促分泌作用及对血细胞的凝集作用.方法采用离子交换色谱法,利用SP Sepharose XL色谱柱纯化重组的免疫调节蛋白;将纯化后的蛋白作用于小鼠脾淋巴细胞和血细胞,用MTT法检测蛋白对脾细胞增殖和血细胞凝集的影响;用ELISA法检测蛋白对淋巴细胞分泌细胞因子的作用.结果经过SP Sepharose XL色谱柱纯化后可获得纯度为86%的重组蛋白;纯化蛋白浓度在80μg/m L时最大限度促进细胞增殖;蛋白浓度在2μg/m L时开始出现凝血;ELISA检测结果显示:重组的金针菇免疫调节蛋白能明显增强小鼠脾淋巴细胞分泌白细胞介素2(IL-2)的水平.结论经过SP Sepharose XL色谱柱纯化后可获得纯度较高的蛋白;纯化后的蛋白与小鼠脾淋巴细胞共同培养,不仅可以促进脾细胞的增殖,而且能增强小鼠脾淋巴细胞分泌白细胞介素2的水平;蛋白与血细胞共同培养能够促进血细胞的凝集.

On Purification and Biological Activity of Recombinant Flammulina Velutipes Immunomodulatory Protein

Objective To study the effects of reFIP-fve on spleen lymphocytes proliferation,interleukin-2( IL-2) secretion from spleen lymphocytes,and on the blood cells agglutination in mice. Method The reFIP-fve was purified through ion exchange chromatography, by using SP Sepharose XL column. MTT method was used to detect the effects of the purified protein on spleen cells proliferation and blood cells agglutination. And the function of reFIP-fve in cytokine secretion from lymphocytes was tested by ELISA method. Results A purity of 86% recombinant protein could be obtained by using SP Sepharose XL column. The purified protein could enhance maximum cell proliferation in the concentration of 80μg/mL,and clotting appeared in the concentration of 2 μg/mL. ELISA showed that the protein significantly enhanced the secretion level of IL-2 from spleen lymphocytes in mice. Conclusion A high purity protein could be obtained through SP Sepharose XL column. When co-cultured with spleen lymphocytes, the purified protein could promote spleen cells proliferation, and enhance the secretion of IL-2 as well. The protein co-cultured with blood cells can accelerate blood cell agglutination.