ENOD40B（一种植物肽激素）的重组生产、同位素标记及纯化
Recombinant production, isotope labeling and purification of ENOD40B: a plant peptide hormone.
作者信息
Chae Young Kee, Tonneli Marco, Markley John L
机构信息
Department of Chemistry and Institute for Chemical Biology, Sejong University, Seoul 143-747, Korea.
出版信息
Protein Pept Lett. 2012 Aug;19(8):808-11. doi: 10.2174/092986612801619624.
Abstract
The plant peptide hormone ENOD40B was produced in a protein production strain of Escherichia coli harboring an induction controller plasmid (Rosetta(DE3)pLysS) as a His6-tagged ubiquitin fusion protein. The fusion protein product was denatured and refolded as part of the isolation procedure and purified by immobilized metal ion chromatography. The peptide hormone was released from its fusion partner by adding yeast ubiquitin hydrolase (YUH) and subsequently purified by reversed phase chromatography. The purity of the resulting peptide fragment was assayed by MALDITOF mass spectrometry and NMR spectroscopy. The final yields of the target peptide were 7.0 mg per liter of LB medium and 3.4 mg per liter of minimal medium.
摘要
植物肽激素ENOD40B在携带诱导控制质粒（Rosetta(DE3)pLysS）的大肠杆菌蛋白质生产菌株中作为His6标记的泛素融合蛋白产生。融合蛋白产物在分离过程中进行变性和复性，并通过固定化金属离子色谱法纯化。通过添加酵母泛素水解酶（YUH）从其融合伴侣中释放出肽激素，随后通过反相色谱法纯化。通过基质辅助激光解吸电离飞行时间质谱（MALDITOF）和核磁共振光谱（NMR）对所得肽片段的纯度进行测定。目标肽的最终产量为每升LB培养基7.0毫克，每升基本培养基3.4毫克。
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