猪链球菌2型溶血素的纯化及生物学活性研究

Purification and biological activities analysis of Streptococcus suis Serotype 2 suilysin

目的:摸索天然和重组猪链球菌2型溶血素的纯化工艺,评价制备蛋白的生物学活性。方法:天然猪溶血素的纯化采用硫酸铵沉淀,阴离子交换层析和疏水层析制备,重组猪溶血素采用镍柱亲和层析进行柱上变复性后,用Th iolproyl-sepharose 6B进一步亲和纯化。通过溶血实验,细胞毒性测定实验评价纯化蛋白的活性。结果:制备的天然和重组猪溶血素纯度均在90%以上,具有较高的溶血活性,较高浓度时能损伤靶细胞,胆固醇能够完全封闭其活性。结论:制备获得的重组猪溶素与天然蛋白具有相近的生物学活性,为进一步研究猪链球菌2型溶血素在猪链球菌致病机制方面的作用奠定了基础。

AIM：To explore the purification methods of wild-type and recombinant suilysin and to evaluate their biological activities.METHODS： Wild-type suilysin was purified by ammonium sulfate precipitation,anion-exchange chromatography and hydrophobic chromatography in turn,while recombinant suilysin was first refolded and purified by immobilized metal ion affinity chromatography,and further purified by Thiopropyl Sepharose 6B.The biological activities were evaluated by hemolysis test,cytotoxicity assay.RESULTS： Both prepared wild-type and recombinant suilysin,with purify over 90%,have hemolysis activity and could injure target cells at high concentration while cholesterol could completely inhibit their activities.CONCLUSION： Recombinant suilysin has similar biological activities with wild-type suilysin,and this work contributed to further study the functions of suilysin on pathogenesis of steptococcus suis.