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頁籤選單縮合
題名 | Expression and Characterization of Rice Manganese Superoxide Dismutase in Escherichia Coli=水稻超氧歧化酶在大腸桿菌中的表現及生化性質研究 |
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作者 | 曾尹貞; 潘素美; Tzeng, Yin-chen; Pan, Shu-mei; |
期刊 | Taiwania |
出版日期 | 19991200 |
卷期 | 44:4 1999.12[民88.12] |
頁次 | 頁479-490 |
分類號 | 434.113 |
語文 | eng |
關鍵詞 | 超氧歧化酶; 含錳超氧歧化酶; 水稻; 大腸桿菌; Superoxide dismutase; SOD; MnSOD; Oryza sativa; E coli; |
中文摘要 | 本文報告水稻超氧歧化 (MnSOD)在大腸桿菌中的表現及生化性質的分析。利用PCR將對應到水稻MnSOD成熟蛋白質之正確cDNA序列接入pGEX-4T-1表現載體中,將建構好的質體再轉型至大腸桿菌BL21品系中。融合蛋白質GST-MnSOD可被IPTG誘導表現,而經由glutathione親和膠體層析可純化均質之GST-MnSOD融合蛋白質。重組之MnSOD(rMnSOD)及GST-MnSOD均對KCN和H C 不敏感,此即保有生物中原態MnSOD的一特性。兩者的單體分子量分別為23 kDa和50 kDa,而在原態上均以二元體的形式存生。rMnSOD的等電點為4.64,GST-MnSOD的等電點則介於pH4.74-4.975之間。rMnSOD及GST-MnSOD兩蛋白質處理60℃,20分鐘後,SOD活性下降至30%;兩者在鹼性環境中均較穩定。 |
英文摘要 | The corrected cDNA coding for rice mature MnSOD protein was made by PCR and inserted into pGEX-4T-1 expression vector. The recombinant DNA was transformed to E. coli BL21 and expression of GST-MnSOD fusion protein was induced by addition of IPTG to bacterial cultures. The homogeneous GST-MnSOD was purified by the one-step GST-glutathione affinity system. Both purified GST-MnSOD and recombinant MnSOD (rMnSOD) remained MnSOD activity, which showed an insensitivity towards KCN and H O . The molecular size of monomer of GST-MnSOD or rMnSOD was 50 kDa and 23 kDa respectively, and the functional form of both was dimer. The isoelectric point of rMnSOD was 4.64, but that of GST-MnSOD was in the range of pH 4.74-4.97. The SOD activities of GST-MnSOD and rMnSOD declined to 30% after incubation at 60℃ for 20 minutes; but they were more stable in an alkaline pH environment. |
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