High hydrostatic pressure enables almost 100% refolding of recombinant human ciliary neurotrophic factor from inclusion bodies at high concentration

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	High hydrostatic pressure enables almost 100% refolding of recombinant human ciliary neurotrophic factor from inclusion bodies at high concentration
	Wang, Qi 1,2; Liu, Yongdong 1; Zhang, Chun 1; Guo, Fangxia 1,2; Feng, Cui 1; Li, Xiunan 1; Shi, Hong 1; Su, Zhiguo 1
刊名	PROTEIN EXPRESSION AND PURIFICATION
	2017-05-01
卷号	133 页码:152-159
关键词	rhCNTF High hydrostatic pressure Inclusion bodies Hydrophobic aggregation Refolding
ISSN号	1046-5928
英文摘要	Protein refolding from inclusion bodies (IBs) often encounters a problem of low recovery at high protein concentration. In this study, we demonstrated that high hydrostatic pressure (HHP) could simultaneously achieve high refolding concentration and high refolding yield for IBs of recombinant human ciliary neurotrophic factor (rhCNTF), a potential therapeutic for neurodegenerative diseases. The use of dilution refolding obtained 18% recovery at 3 mg/mL, even in the presence of 4 M urea. In contrast, HHP refolding could efficiently increase the recovery up to almost 100% even at 4 mg/mL. It was found that in the dilution, hydrophobic aggregates were the off-path products and their amount increased with the protein concentration. However, HHP could effectively minimize the formation of hydrophobic aggregates, leading to almost complete conversion of the rhCNTF IBs to the correct configuration. The stable operation range of concentration is 0.5-4.0 mg/mL, in which the refolding yield was almost 100%. Compared with the literatures where HHP failed to increase the refolding yield beyond 90%, the reason could be attributed to the structural difference that rhCNTF has no disulfide bond and is a monomeric protein. After purification by one-step of anionic chromatography, the purity of rhCNTF reached 95% with total process recovery of 54.1%. The purified rhCNTF showed similar structure and in vitro bioactivity to the native species. The whole process featured integration of solubilization/refolding, a high refolding yield of 100%, a high concentration of 4 mg/mL, and a simple chromatography to ensure a high productivity. (C) 2017 Elsevier Inc. All rights reserved.
WOS标题词	Science & Technology ; Life Sciences & Biomedicine
类目[WOS]	Biochemical Research Methods ; Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology
研究领域[WOS]	Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology
关键词[WOS]	FACTOR CNTF ; ESCHERICHIA-COLI ; GROWTH-HORMONE ; PROTEINS ; PURIFICATION ; EXPRESSION ; COMPRESSIBILITY ; SOLUBILIZATION ; OPTIMIZATION ; WEIGHT
收录类别	SCI
语种	英语
WOS记录号	WOS:000401684400019
内容类型	期刊论文
源URL	[http://ir.ipe.ac.cn/handle/122111/22544]
专题	过程工程研究所_生化工程国家重点实验室
作者单位	1.Chinese Acad Sci, Natl Key Lab Biochem Engn, Inst Proc Engn, 1 Beierjie St, Beijing 100190, Peoples R China 2.Univ Chinese Acad Sci, Beijing 100049, Peoples R China
推荐引用方式 GB/T 7714	Wang, Qi,Liu, Yongdong,Zhang, Chun,et al. High hydrostatic pressure enables almost 100% refolding of recombinant human ciliary neurotrophic factor from inclusion bodies at high concentration[J]. PROTEIN EXPRESSION AND PURIFICATION,2017,133:152-159.
APA	Wang, Qi.,Liu, Yongdong.,Zhang, Chun.,Guo, Fangxia.,Feng, Cui.,...&Su, Zhiguo.(2017).High hydrostatic pressure enables almost 100% refolding of recombinant human ciliary neurotrophic factor from inclusion bodies at high concentration.PROTEIN EXPRESSION AND PURIFICATION,133,152-159.
MLA	Wang, Qi,et al."High hydrostatic pressure enables almost 100% refolding of recombinant human ciliary neurotrophic factor from inclusion bodies at high concentration".PROTEIN EXPRESSION AND PURIFICATION 133(2017):152-159.