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学科主题: 天然产物有机化学
题名: The expression and refolding of isotopically labeled recombinant Matrilysin for NMR studies overexpression; purification; refolding
其他题名: 用于NMR 研究的Matrilysin同位素标记重组子的表达和重折叠
作者: Ou L(欧力) ; Ma JB(麻锦彪) ; Zheng XH(郑训海) ; Chen X(陈翔) ; Li GY(李光耀) ; Wu HM(吴厚铭)
通讯作者: 吴厚铭
刊名: Protein Expr. Purif.
发表日期: 2006-01-01
卷: 47, 期:2, 页:367-373
收录类别: SCI
部门归属: 上海有机化学研究所生命有机化学国家重点实验室
英文摘要: Matrilysin (MMP7) is the smallest member of matrix metalloproteinases (MMPs) family, which are collectively responsible for remodeling of connective tissue. MMP7 plays an essential role in cancer, innate immunity, and in inflammatory disorders, and has been justified as a novel drug target. Here, We report the gene synthesis, overexpression in Escherichia coli, purification and refolding of MMP7. The gene of Matrilysin was synthesized based on PCR method and overexpressed in E. coli in the form of inclusion bodies. The protein was subsequently purified and refolded to yield sufficient quantities for structural and functional studies. The purified protein was characterized by means of MALDI-TOF mass spectroscopy and dynamic light scattering (DLS) analysis. The MS data confirms the correctness of the primary sequence, while DLS experiment proves that the protein exists as a monomeric form. A significantly optimized protocol has been worked out to prepare N-15 and/or C-13-labeled MMP7 in minimal medium with high yields for NMR studies. Under the various conditions optimized for the purification of MMP7, the yield of the purified protein is estimated to be 18-20 mg from 0.5 L of M9 minimal media. Finally, the N-15-1H HSQC spectrum of uniformly N-15-labeled MMP7 sample reveals that the protein is properly folded, and exists in a well-ordered structure. (c) 2006 Elsevier Inc. All rights reserved.
语种: 英语
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内容类型: 期刊论文
URI标识: http://ir.sioc.ac.cn/handle/331003/17369
Appears in Collections:上海有机化学研究所_期刊论文

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Recommended Citation:
Ou L,Ma JB,Zheng XH,et al. The expression and refolding of isotopically labeled recombinant Matrilysin for NMR studies overexpression; purification; refolding[J]. Protein Expr. Purif.,2006,47(2):367-373.
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