Identification, characterization, and stabilization of the deamidation degradation of recombinant human tumor necrosis factor-alpha
文献类型：期刊论文
| 作者 | Yin, Shuang1,2; Zhang, Chun1,2; Li, Zenglan2; Wang, Qi2; Shi, Hong2; Yu, Rong1; Liu, Yongdong2; Su, Zhiguo2 |
| 刊名 | PROCESS BIOCHEMISTRY
 |
| 出版日期 | 2017-02-01 |
| 卷号 | 53页码:216-223 |
| 关键词 | Deamidation Tumor necrosis factor-alpha LC-MS/MS Polyol Purification |
| ISSN号 | 1359-5113 |
| 英文摘要 | A kind of degradation characterized by an increase in overall negative charge in both native polyacrylamide gel electrophoresis analysis and high-performance strong anion exchange analysis was observed during the purification process of recombinant human tumor necrosis factor-alpha (TNF-alpha). Liquid chromatography coupled with tandem mass spectrometry was adopted to further analyze this degradation, and the result demonstrated that suspected deamidation occurred at N39 and N34 residues. To investigate the effects of these deamidation degradations on TNF-alpha, we substituted corresponding asparagine residues with aspartic acid residues. High-performance size-exclusion chromatography, circular dichroism, and fluorescence spectrometry analysis revealed that the advanced structures of TNF-alpha could not be obviously changed by these substitutions. Differential scanning calorimetry analysis indicated that deamidation led to decreased thermal stability, and two mutants (N34D, N34DN39D) both possessed two Tm. L929 cell cytotoxic activity implied that N39 residue deamidation caused only a minor bioactivity loss, whereas N34 residue deamidation led to a bioactivity loss of four orders of magnitude. To alleviate the degradation during the purification process, we screened nine excipients and found that glycerol could notably ameliorate this degradation and provide a compromise strategy for the recombinant human TNF-alpha protein during purification process and formulation development. (C) 2016 Published by Elsevier Ltd. |
| WOS标题词 | Science & Technology ; Life Sciences & Biomedicine ; Technology |
| 类目[WOS] | Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology ; Engineering, Chemical |
| 研究领域[WOS] | Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology ; Engineering |
| 关键词[WOS] | PURIFICATION ; ANTIBODIES ; PROTEINS ; ASPARTYL ; ANTIGEN |
| 收录类别 | SCI |
| 语种 | 英语 |
| WOS记录号 | WOS:000394403400026 |
| 源URL | [http://ir.ipe.ac.cn/handle/122111/21989]  |
| 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.Sichuan Univ, West China Sch Pharm, Minist Educ, Key Lab Drug Targeting & Drug Delivery Syst, Chengdu 610041, Peoples R China 2.Chinese Acad Sci, Inst Proc Engn, State Key Lab Biochem Engn, Beijing 100080, Peoples R China |
| 推荐引用方式 GB/T 7714 | Yin, Shuang,Zhang, Chun,Li, Zenglan,et al. Identification, characterization, and stabilization of the deamidation degradation of recombinant human tumor necrosis factor-alpha[J]. PROCESS BIOCHEMISTRY,2017,53:216-223. |
| APA | Yin, Shuang.,Zhang, Chun.,Li, Zenglan.,Wang, Qi.,Shi, Hong.,...&Su, Zhiguo.(2017).Identification, characterization, and stabilization of the deamidation degradation of recombinant human tumor necrosis factor-alpha.PROCESS BIOCHEMISTRY,53,216-223. |
| MLA | Yin, Shuang,et al."Identification, characterization, and stabilization of the deamidation degradation of recombinant human tumor necrosis factor-alpha".PROCESS BIOCHEMISTRY 53(2017):216-223. |
入库方式： OAI收割
来源：过程工程研究所
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