Hydrophobic interaction chromatography correctly refolding proteins assisted by glycerol and urea gradients
文献类型：期刊论文
| 作者 | Li, JJ; Liu, YD; Wang, FW; Ma, GH; Su, ZG |
| 刊名 | JOURNAL OF CHROMATOGRAPHY A
 |
| 出版日期 | 2004-12-24 |
| 卷号 | 1061期号:2页码:193-199 |
| 关键词 | refolding hydrophobic interaction chromatography glycerol lysozyme gradient |
| ISSN号 | 0021-9673 |
| 其他题名 | J. Chromatogr. A |
| 中文摘要 | Chromatographic columns packed with commercially available hydrophobic interaction chromatography (HIC) media were found to be able to suppress aggregation and nevertheless had a tendency to promote the structural misfolding resulting in higher soluble protein recovery and lower specific activity than that by dilution when they were used to refold lysozyme. a model protein. Moreover, this misfolding effect was exacerbated with increasing hydrophobicity of media. A novel strategy involving the combination of glycerol, a typical osmolyte, a urea gradient and commercially available HIC media was introduced to facilitate protein refolding correctly as well as improve mass recovery by providing a gradual change of the refolding environment in the HIC column. In this process, unfolded lysozyme was bound to Poros PE HIC column at high salt concentration and was released by a urea gradient followed by elution with refolding buffer in the presence of 50% (v/v) glycerol, resulting in 86.3% activity yield and 85% mass recovery with the refolded product of native specific activity For the absence of glycerol, only 50.9% activity yield and 59% specific activity recovery was obtained although mass recovery was closed to that in the presence of glycerol. It was also discovered that glycerol addition during elution process was necessary for correct refolding compared to mixing of glycerol with post-column fraction. The possible mechanism for refolding with this system vas proposed to be relevant to the formation of an on-pathway intermediate that could slowly reactivate. (C) 2004 Elsevier B.V. All rights reserved. |
| 英文摘要 | Chromatographic columns packed with commercially available hydrophobic interaction chromatography (HIC) media were found to be able to suppress aggregation and nevertheless had a tendency to promote the structural misfolding resulting in higher soluble protein recovery and lower specific activity than that by dilution when they were used to refold lysozyme. a model protein. Moreover, this misfolding effect was exacerbated with increasing hydrophobicity of media. A novel strategy involving the combination of glycerol, a typical osmolyte, a urea gradient and commercially available HIC media was introduced to facilitate protein refolding correctly as well as improve mass recovery by providing a gradual change of the refolding environment in the HIC column. In this process, unfolded lysozyme was bound to Poros PE HIC column at high salt concentration and was released by a urea gradient followed by elution with refolding buffer in the presence of 50% (v/v) glycerol, resulting in 86.3% activity yield and 85% mass recovery with the refolded product of native specific activity For the absence of glycerol, only 50.9% activity yield and 59% specific activity recovery was obtained although mass recovery was closed to that in the presence of glycerol. It was also discovered that glycerol addition during elution process was necessary for correct refolding compared to mixing of glycerol with post-column fraction. The possible mechanism for refolding with this system vas proposed to be relevant to the formation of an on-pathway intermediate that could slowly reactivate. (C) 2004 Elsevier B.V. All rights reserved. |
| WOS标题词 | Science & Technology ; Life Sciences & Biomedicine ; Physical Sciences |
| 类目[WOS] | Biochemical Research Methods ; Chemistry, Analytical |
| 研究领域[WOS] | Biochemistry & Molecular Biology ; Chemistry |
| 关键词[WOS] | SIZE-EXCLUSION CHROMATOGRAPHY ; INCLUSION-BODIES ; ESCHERICHIA-COLI ; LYSOZYME ; RENATURATION ; AGGREGATION ; YIELD |
| 收录类别 | SCI |
| 原文出处 | |
| 语种 | 英语 |
| WOS记录号 | WOS:000225853400009 |
| 公开日期 | 2013-11-05 |
| 版本 | 出版稿 |
| 源URL | [http://ir.ipe.ac.cn/handle/122111/4944]  |
| 专题 | 过程工程研究所_研究所（批量导入） |
| 作者单位 | 1.Chinese Acad Sci, Inst Proc Engn, Natl Key Lab Biochem Engn, Beijing 100080, Peoples R China 2.Dalian Univ Technol, Dalian 116024, LiaoNing, Peoples R China 3.Dalian Media Univ, Clin Coll 2, Dalian 116027, LiaoNing, Peoples R China |
| 推荐引用方式 GB/T 7714 | Li, JJ,Liu, YD,Wang, FW,et al. Hydrophobic interaction chromatography correctly refolding proteins assisted by glycerol and urea gradients[J]. JOURNAL OF CHROMATOGRAPHY A,2004,1061(2):193-199. |
| APA | Li, JJ,Liu, YD,Wang, FW,Ma, GH,&Su, ZG.(2004).Hydrophobic interaction chromatography correctly refolding proteins assisted by glycerol and urea gradients.JOURNAL OF CHROMATOGRAPHY A,1061(2),193-199. |
| MLA | Li, JJ,et al."Hydrophobic interaction chromatography correctly refolding proteins assisted by glycerol and urea gradients".JOURNAL OF CHROMATOGRAPHY A 1061.2(2004):193-199. |
入库方式： OAI收割
来源：过程工程研究所
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