High-level expression and purification of Escherichia coli oligopeptidase B
文献类型：期刊论文
| 作者 | Yan, JB; Wang, GQ; Du, P; Zhu, DX; Wang, MW ; Jiang, XY
|
| 刊名 | PROTEIN EXPRESSION AND PURIFICATION
 |
| 出版日期 | 2006-06 |
| 卷号 | 47期号:2页码:645-650 |
| 关键词 | Escherichia coli oligopeptidase B trypsin-like protease fusion expression |
| ISSN号 | 1046-5928 |
| DOI | 10.1016/j.pep.2006.01.018 |
| 文献子类 | Article |
| 英文摘要 | Oligopeptidase B (OpdB) of Escherichia coli, previously called protease 11, has a trypsin-like specificity, cleaving peptides at lysine and arginine residues and belongs to the prolyl oligopeptidase family of new serine peptidases. In this study, we report the fusion expression of E coli oligopeptidase B with an N-terminal histidine tag using pET28a as the expression vector. Although most of the recombinant OpdB was produced as inclusion bodies, the solubility of the recombinant protease increased significantly when the expression temperature shifted from 37 to 30 degrees C. Recombinant OpdB (similar to 10 mg) could be purified from the soluble fraction of the crude extract of 1 L log-phase E. coli culture containing 1.5 g wet bacterial cells. The purified OpdB has a molecular weight of similar to 80 kDa and a specific activity of 4.8 x 10(4) U/Mg. OpdB could also be purified from the inclusion bodies with a lower yield. The recombinant enzyme was very stable under 40 degrees C. By comparison of the substrate specificity of the purified OpdB with that of OpdA, another trypsin-like protease in E coli, we found that Boc-Glu-Lys-Lys-MCA is a specific substrate for E coli OpdB. We also found that compared to OpdA, OpdB is much more sensitive to GMCHA-OPhBu, a synthetic trypsin inhibitor that can retard the growth of E coli. (c) 2006 Elsevier Inc. All rights reserved. |
| WOS关键词 | ACID 4-TERT-BUTYLPHENYL ESTER ; PROTEASE-II ; TRYPSIN-INHIBITORS ; ENZYME ; TYPHIMURIUM ; GROWTH ; GENE |
| WOS研究方向 | Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology |
| 语种 | 英语 |
| WOS记录号 | WOS:000238277000039 |
| 出版者 | ACADEMIC PRESS INC ELSEVIER SCIENCE |
| 源URL | [http://119.78.100.183/handle/2S10ELR8/273585]  |
| 专题 | 中国科学院上海药物研究所 |
| 通讯作者 | Jiang, XY |
| 作者单位 | 1.Nanjing Univ, State Key Lab Pharmaceut Biotechnol, Nanjing 210093, Peoples R China 2.Nanjing Univ, Dept Biochem, Nanjing 210093, Peoples R China 3.Chinese Acad Sci, Shanghai Inst Mat Med, Natl Ctr Drug Screening, Shanghai 201203, Peoples R China |
| 推荐引用方式 GB/T 7714 | Yan, JB,Wang, GQ,Du, P,et al. High-level expression and purification of Escherichia coli oligopeptidase B[J]. PROTEIN EXPRESSION AND PURIFICATION,2006,47(2):645-650. |
| APA | Yan, JB,Wang, GQ,Du, P,Zhu, DX,Wang, MW,&Jiang, XY.(2006).High-level expression and purification of Escherichia coli oligopeptidase B.PROTEIN EXPRESSION AND PURIFICATION,47(2),645-650. |
| MLA | Yan, JB,et al."High-level expression and purification of Escherichia coli oligopeptidase B".PROTEIN EXPRESSION AND PURIFICATION 47.2(2006):645-650. |
入库方式： OAI收割
来源：上海药物研究所
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