实现人源FGF-21高效可溶性表达的两种策略

华东师范大学学报(自然科学版) ›› 20120, Vol. 2012 ›› Issue (6): 114-121.doi:

实现人源FGF-21高效可溶性表达的两种策略

李娟，刘雯，肖磊，劳勋，赵丽芬，黄静，吴自荣

华东师范大学生命科学学院，上海 200062

收稿日期:2011-09-01 修回日期:2011-12-01 出版日期:2012-11-25 发布日期:2013-01-11

Two strategies for efficient expression of soluble recombinant human FGF-21 protein

LI Juan, LIU Wen, XIAO Lei, LAO Xun, ZHAO Li-fen,HUANG Jing, WU Zi-rong

School of Life Sciences, East China Normal University, Shanghai 200062， China

Received:2011-09-01 Revised:2011-12-01 Online:2012-11-25 Published:2013-01-11

摘要/Abstract

摘要： 采用分子伴侣协助蛋白表达的方法及乳糖自诱导的策略，实现人源FGF-21在大肠杆菌中的高效可溶性表达.结果显示，在分子伴侣Tig的协同表达下，融合蛋白TrxA-FGF-21可溶性达到92.4%.采用乳糖自诱导策略，融合蛋白TrxA-FGF-21表达量达到17.4%左右，菌体〖WTBX〗OD〖WTBZ〗600达到12.4，可溶性TrxA-FGF-21相对产量是IPTG诱导发酵条件的5.5倍.该方法制备的重组FGF-21在C57BL/6小鼠体内具有降低血糖的生物学活性，甘油三酯水平与对照组相比也表现极显著差异(〖WTBX〗P〖WTBZ〗<0.01).

关键词: 成纤维生长因子21, 分子伴侣, 乳糖自诱导

Abstract: The expression of recombinant soluble FGF-21 protein was achieved through transferring another plasmid pTf16 which expressed molecular chaperone Tig. On the other hand, the yield of FGF-21 was greatly improved with the method of lactose auto-induction. The result showed that the soluble TrxA-FGF-21 fusion protein reached 92.4% by molecular chaperone co-expression. With lactose auto-induction, the expression quantity of fusion protein was 17.4% of the total proteins, and the 〖WTBX〗OD600 〖WTBZ〗 was 12.4; thus relative yield was 5.5 times higher than IPTG induction. The biological activities of recombinant FGF-21 were studied 〖WTBX〗in vivo〖WTBZ〗 in healthy C57BL/6 mice. It showed that recombinant FGF-21 can lower blood glucose obviously in mice (〖WTBX〗P〖WTBZ〗 <0.05 or 〖WTBX〗P 〖WTBZ〗<0.01). The recombinant FGF-21 treatment also led to a significant decrease in concentration of triglyceride in C57BL/6 mice (〖WTBX〗P〖WTBZ〗 <0.01).

Key words: fibroblast growth factor 21, molecular chaperone, lactose auto-induction

中图分类号:

Q789

李娟，刘雯，肖磊，劳勋，赵丽芬，黄静，吴自荣. 实现人源FGF-21高效可溶性表达的两种策略[J]. 华东师范大学学报(自然科学版), 20120, 2012(6): 114-121.

LI Juan, LIU Wen, XIAO Lei, LAO Xun, ZHAO Li-fen,HUANG Jing, WU Zi-rong. Two strategies for efficient expression of soluble recombinant human FGF-21 protein[J]. Journal of East China Normal University(Natural Sc, 20120, 2012(6): 114-121.

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参考文献

［1］ NISHIMURA T, NAKATAKE Y, KONISHI M, et al. Identification of a nove FGF, FGF-21, preferentially expressed in the liver［J］. Biochim Biophys Acta, 2000, 1492(1): 203-206.

［2］ IZUMIYA Y, BINA H A, OUCHI N, et al. FGF-21 is an Akt-regulated myokine［J］. FEBS Lett, 2008, 582(17): 3805-3810.

［3］ KHARTONENKOV A, SHIYANOVA T L, KOESTER A, et al. FGF-21 as a novel metabolic regulator［J］. J Clin Invest, 2005, 115(6): 1627-1635.

［4］ WENTE W, EFANOV A M, BRENNER M, et al. Fibroblast growth factor-21 improves pancreatic beta-cell function and survival by activation of extracellular signal-regulated kinase 1/2 and Akt signaling pathways［J］. Diabetes, 2006, 55(9): 2470-2478.

［5］ COSKUN T, BINA H A, SCHNEIDER M A, et al. Fibroblast growth factor 21 corrects obesity in mice［J］. Endocrinology, 2008, 149(12): 6018-6027.

［6］姜媛媛，尹成凯，李晋南，等.SUMO 融合系统高效表达可溶性重组蛋白的研究［J］.东北农业大学学报，2008, 39(10): 57-62.

［7］崔硕硕，林学政，沈继红.分子伴侣共表达对低温脂肪酶Lip-837异源可溶性表达的影响［J］.海洋科学进展，2011, 29(1):105-111.

［8］ ARAYO H, KAZUNORI H, NOBUTADA T, et al. Scalable purification and characterization of the extracellular domain of human autotaxin from prokaryotic cells［J］. Protein Expression and Purification, 2008, 59(1): 9-17.

［9］顾娟，劳勋，金明飞，等.人胰高血糖素样肽-1突变体融合蛋白在大肠杆菌中的自诱导表达优化［J］.微生物学通报，2010, 37(5): 726-731.

［10］冯杉.T7表达系统及自诱导蛋白产出策略［J］.北京教育学院学报:自然科学版，2009, 4(3): 10-15. 

［11］ COFFEY R J Jr, DERYNCK R, WILCOX J N, et al. Production and auto-induction of transforming growth factor-alpha in human keratinocyte［J］. Nature, 1987, 328(6133): 817-820.

［12］ STUDIER F W. Protein production by auto-induction in high density shaking cultures［J］. Protein Expr Purif, 2005, 41(1): 207-234.

［13］ LU Z S, CHEN W T, DING Y, et al. A novel method for high-level production of psychrophilic TAB5 alkaline phosphatase［J］. Protein Expression and Purification, 2010, 74(2): 217-222.

［14］刘爽，胡宝成.原核系统可溶性表达策略［J］.生物技术通讯，2005(2): 172-175.