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目的表达纯化不可分型流感嗜血杆菌表面天冬氨酸酶(r ASP)和其敲除羧基末端赖氨酸的重组蛋白(r ASPΔK)。方法克隆了不可分型流感嗜血杆菌ATCC49247的ASP基因和ASPΔK基因,与载体连接后,转入大肠杆菌表达蛋白,并用Co2+亲和柱层析纯化蛋白;采用酶促反应测定纯化蛋白的酶活性。结果两种基因的克隆和表达质粒构建成功,目的蛋白表达量大且具有较高的酶活性。结论成功表达纯化了r ASP和r ASPΔK蛋白。
Objective To express and purify untranslatable Haemophilus influenzae surface aspartase (r ASP) and its recombinant protein (r ASPΔK) which knocked out carboxyl-terminal lysine. Methods The ASP gene and ASPΔK gene of Haemophilus influenzae type ATCC 49247 were cloned. After being linked with the vector, the recombinant protein was transferred into E. coli and the protein was purified by Co2 + affinity column chromatography. The enzymatic activity of purified protein was determined by enzymatic reaction. Results The cloning and expression plasmids of the two genes were successfully constructed. The target protein was expressed in a large amount and had high enzyme activity. Conclusion The r ASP and r ASPΔK proteins were successfully expressed and purified.