摘要
目的复性黄牛朊蛋白原核表达产物,分析复性蛋白的活性,以获得有活性的牛重组朊蛋白。方法构建并诱导表达黄牛朊蛋白基因重组菌,复性其包涵体裂解物,对GST-BoPrP(93～241)(Q234R)复性产物进行GSTrap FF柱层析纯化,通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳和免疫印迹观察其提纯和复性效果。结果重组菌可表达分子质量约为42.4 ku的目的蛋白,包涵体裂解物经过透析复性后,可获得较高纯度的复性目的蛋白。各复性产物和纯化的GST-BoPrP(93～241)(Q234R)蛋白经免疫印迹鉴定,发现仅分子质量约为42.4 ku的条带与单抗4C11结合。结论获得高纯度复性黄牛朊蛋白原核表达蛋白,具有良好的免疫反应活性。
Objective To renature the bovine prion protein with prokaryotic expression,analyze their immune response activity,and to obtain the active recombinant bovine PrPs in final. Methods Bovine prion protein gene recombinant bacteria were constructed and expressed,and the products were renatured.GST-BoPrP（93～241）（Q234R） was purified through GSTrap FF column and identified by sodium dodecyl sulfonate polyacrylamide gel electrophoresis and western blotting.Results The protein about 42.4ku was expressed by recombinant E.coli and high-purity protein was obtained after the inclusion bodies denatured and renatured.The expectant molecular about 42.4 ku of all the renatured products and purified GST-BoPrP（93～241） （Q234R） could response to the anti-bovine PrP monoclonal antibody 4C11.Conclusion Highpurity renatured protein of bovine PrPs prokaryotic expression is obtained and has a good immune response activity.
出处
《兰州大学学报（医学版）》
CAS
2010年第4期16-20,共5页
Journal of Lanzhou University（Medical Sciences）
基金
高校博士点专项科研基金(20060733006)
家畜疫病病原生物学国家重点实验室基金(2008-2009)
关键词
牛朊蛋白
原核表达
复性
活性分析
bovine prion protein
prokaryotic expression
renaturation
activity determination
