| His-tag标记的嗜热菌来源L-脯氨酸脱氢酶的纯化与性质 |
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| 引用本文： | 林乐懿,张丽叶,末信一郎,郑海涛,白石智美,桶崎陽友.His-tag标记的嗜热菌来源L-脯氨酸脱氢酶的纯化与性质[J].北京化工大学学报(自然科学版),2010,37(6):121-125. |
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| 作者姓名： | 林乐懿 张丽叶 末信一郎 郑海涛 白石智美 桶崎陽友 |
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| 作者单位： | 1北京化工大学 生命科学与技术学院， 北京 100029; 2日本福井大学 生物应用化学部， 日本 福井 9108507;3天津工业大学 环境与化学工程学院， 天津 300160 |
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| 摘 要： | 以引入了His-tag做标记的超嗜热菌Thermococcus profundus色素依存性L-脯氨酸脱氢酶基因片段为目的基因,在宿主大肠杆菌Rosetta-gami(DE3)CompetentCells中表达目的蛋白。细胞培养液经超声菌体破碎、热处理、NiSepharose层析分离,得到纯度较高的His-tag色素依存性L-脯氨酸脱氢酶,比酶活是纯化前的5倍。纯化后的酶液经凝胶电泳分离得到片段为58600,40200,22300的3段短肽。通过对该酶性质的初步探索,得出在温度50℃,体系pH8.0,300mmol/LTris缓冲溶液的条件下,L-脯氨酸脱氢酶酶活最高,为1.5U/mL。
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| 关 键 词： | 超嗜热菌 His-tag 色素依存性L-脯氨酸脱氢酶 镍亲和色谱 |
| 收稿时间： | 2010-04-14 |
Purification and characterization of His-tag L-proline dehydrogenase from a hyperthermophilic archaeon, Thermococcus profundus |
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| LIN LeYi,ZHANG LiYe,Shin-ichiro SUYE,ZHENG HaiTao,Satomi SHIRAISHI,Yosuke OKEZAKI.Purification and characterization of His-tag L-proline dehydrogenase from a hyperthermophilic archaeon, Thermococcus profundus[J].Journal of Beijing University of Chemical Technology,2010,37(6):121-125. |
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| Authors: | LIN LeYi ZHANG LiYe Shin-ichiro SUYE ZHENG HaiTao Satomi SHIRAISHI Yosuke OKEZAKI |
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| Institution: | 1. College of Life Science and Technology, Beijing University of Chemical Technology, Beijing 100029, China; 2. Department of Applied Chemistry and Biotechnology, University of Fukui, Fukui 9108507, Japan; 3. School of Environmental and Chemical Engineering, Tianjin Polytechnic University, Tianjin 300160, China |
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| Abstract: | A His-tag marked gene fragment of dye-linked L-proline dehydrogenase (dye-linked L-pro DH) from a hyperthermophilic archaeon,Thermococcus profundus,has been used as the target gene to express the desired protein with Escherichia coli Rosetta-gami (DE3) competent cells. The His-tag L-pro DH was gained through fermentation,and purified by using cell-free extraction,heat treatment and Ni Sepharose chromatography,resulting in a five-fold increase in specific activity. Three peptides with molecular weights of 58... |
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| Keywords: | Hyperthermophilic archaeon His tag dye-linked L-proline dehydrogenase Ni Sepharose |
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