Efficient Expression of Bioactive Human Leptin in <Emphasis Type="Italic">Escherichia coli</Emphasis> in Soluble Fusion Form |
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| Authors: | Jian Feng Li Jie Zhang Zhen Zhang Yun Long Hu Shuang Quan Zhang |
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| Institution: | (1) Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, Life Science College, Nanjing Normal University, Nanjing, 210046, Jiangsu, People’s Republic of China;(2) Jiangsu Engineering Research Center for Biomedical Function Materials, Nanjing Normal University, Nanjing, 210046, Jiangsu, People’s Republic of China;(3) Jiangsu Key Laboratory for Supermolecular Medicinal Materials and Applications, Nanjing Normal University, Nanjing, 210046, Jiangsu, People’s Republic of China; |
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| Abstract: | Leptin, a 16 kDa nonglycosylated hormone, is produced by mature adipocytes and functions primarily in the hypothalamus to
reduce food intake and body weight. To explore a new approach for high-level expression of human Leptin in Escherichia coli, the human Leptin gene, synthesized according to the published sequence, was cloned into the vector pET32a to construct a
fusion expression plasmid: Trx–Leptin/pET32a. Our data showed that more than 40% of the fusion protein Trx–Leptin was expressed
in soluble form. After purified by Ni-IDA affinity chromatography, cleaved by enterokinase and applied Ni-IDA affinity chromatography
again, purified Leptin with homogeneity over 96% was achieved. The bio-functional experiments of purified Leptin showed a
significant reduction in food intake and body weight of female mice treated with Leptin by comparing with control mice, and
it indicated that the purified Leptin has full biological activity. In addition, our expression system was a very low-cost
and efficient prokaryotic expression system. So taken together, our results demonstrated that our expression system of bio-active
Leptin provided a new method for producing Leptin in big scale and would be widely applied in commercial Leptin producing
industries. |
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| Keywords: | Human leptin Trx Escherichia coli Fusion expression Enterokinase Affinity purification |
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