| 蜂毒肽融合蛋白在裂殖酵母中的表达研究 |
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| 引用本文： | 胡宗利,刘志昭,邓磊,潘宇,陈国平.蜂毒肽融合蛋白在裂殖酵母中的表达研究[J].药物生物技术,2010(1). |
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| 作者姓名： | 胡宗利 刘志昭 邓磊 潘宇 陈国平 |
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| 作者单位： | 重庆大学生物工程学院; |
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| 基金项目： | 国家“863”计划项目(2006AA02Z138); 国家自然科学基金项目(No.30771464,30871709)资助 |
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| 摘 要： | 将抗菌肽之一的蜂毒溶血肽(Melittin)基因序列定向克隆到表达载体pESP-2中,构建GST-Melittin融合蛋白。利用载体上严格受硫胺素浓度调控的启动子Pnmtl,采用先扩大培养后诱导表达的两步培养法,实现了融合蛋白在裂殖酵母Schizosaccharomyces Pombe中的异源表达。然后通过GST亲和层析法一步纯化得到融合肽。研究中1L培养液收集到约5g鲜活酵母菌,纯化得到400μg融合蛋白,融合蛋白经SDS-PAGE和Western blot验证,表明蜂毒肽融合蛋白已在裂殖酵母中高效的表达,并可以通过GST亲和层析法得到有效纯化。初步的活性实验证明所获得Melittin蛋白对Escherichia Coli DH5α菌种有良好的抑菌效果。
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| 关 键 词： | 蜂毒肽 裂殖酵母 蛋白表达 GST亲和层析 |
GST-Melittin Expression in Schizosaccharomyces Pombe |
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| HU Zong-li,LIU Zhi-zhao,DENG Lei,PAN Yu,CHEN Guo-ping.GST-Melittin Expression in Schizosaccharomyces Pombe[J].Pharmaceutical Biotechnology,2010(1). |
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| Authors: | HU Zong-li LIU Zhi-zhao DENG Lei PAN Yu CHEN Guo-ping |
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| Institution: | HU Zong-li,LIU Zhi-zhao,DENG Lei,PAN Yu,CHEN Guo-ping(Bioengineering School,Chongqing University,Chongqing 400044,China) |
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| Abstract: | Melittin gene was directionally cloned into pESP-2 expression vector to form a hybrid gene that can produce GST-melittin fusion protein. The heterologous expression of fusion protein in Schizosaccharomyces Pombe was driven by the nmtl promoter (Pnmtl) in pESP-2, which was tightly regulated by thiamine. The two-step culture system was performed and recombinants were first expanded in YES medium containing thiamine; in the second step GST-melittin fusion protein expression was induced in EMM medium without th... |
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| Keywords: | Melittin Schizosaccharomyces Pombe Protein expression GST-affinity chromatography |
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