重组蛋白GST-OsCAT_B的表达特性研究(英文)

Characteristics of Recombined Protein GST-OsCAT_B Expressed in E.coli

为了阐明水稻Catalase(CAT)的酶学功能,首先需要获得出足量的、活性的该酶蛋白。本研究克隆了水稻OsCATB基因(GenBank accession No.D26484),构建到原核表达载体pGEX-6p-3中形成重组蛋白,继而转入E.coli菌株BL21中进行表达特性研究。结果表明,GST-OsCATB融合蛋白在E.coli中进行了过量表达,表达受到诱导剂浓度、诱导时间、诱导温度和诱导体系等多因素影响;通过谷胱甘肽Sepharose-4B亲合层析,纯化出足量、活性的融合蛋白GST-OsCATB,每克表达细胞(干重)中得率为51 mg GST-OsCATB。

To clarify the enzymology of catalase in rice（Oryza sativa L.）,large quantities and active of the target protein is available firstly.Here,rice OsCATB（GenBank accession No.D26484） gene was cloned and constructed with the pGEX-6p-3 vector to allow expression of OsCATB as glutathione-S-transferase（GST） fusion protein,and E.coli strain BL21 was used for expression of fusion proteins as well.The results indicated that GST-OsCATB fusion proteins were effectively expressed in E.coli,and regulated by IPTG,inducing period,temperature and others of the actions.The purified,enough and activity GST-OsCATB was obtained by affinity chromatography using glutathione-Sepharose 4B column.The final yield was 51 mg·g-1 dry cells for GST-OsCATB.