Expression and purification of lipoprotein-associated phospholipase A(2), a key enzyme involved in atherosclerosis
文献类型：期刊论文
| 作者 | Zhang, Fu-Jun; Cai, Mao-Jun; Shen, Jing-Kang; Wang, Yi-Ping
|
| 刊名 | ACTA PHARMACOLOGICA SINICA
 |
| 出版日期 | 2006-06 |
| 卷号 | 27期号:6页码:679-684 |
| 关键词 | lipoprotein-associated phospholipase A2 atherosclerosis coronary heart diseases cloning baculovirus purification |
| ISSN号 | 1671-4083 |
| DOI | 10.1111/j.1745-7254.2006.00308.x |
| 文献子类 | Article |
| 英文摘要 | Aim: To express and purify lipoprotein-associated phospholipase A2 (Lp-PLA(2)), and to establish a screening model for Lp-PLA(2) inhibitors using the expressed Lp-PLA(2). Methods: We cloned the full-length cDNA of Lp-PLA(2) from differentiated THP-1 cells, and subcloned the cDNA into the baculovirus transfer vector pFastBac1. In addition, we introduced an N-terminal Kozak sequence for high-level translation initiation and a C-terminal sequence of 6 histidine residues for purification. The fusion enzyme was expressed in Sf9 insect cells and purified by Ni2+ affinity chromatography. Recombinant Lp-PLA(2) activity was measured using [H-3]PAF as a substrate, and we examined the enzyme activity of recombinant Lp-PLA(2) pretreated with the known Lp-PLA(2) inhibitor SB435495. Results: We successfully cloned the full-length Lp-PLA(2) gene from differentiated THP-1 cells. The fusion enzyme was expressed in Sf9 insect cells at a high level and purified efficiently through a 2-step procedure. The recombinant Lp-PLA(2) activity was measured using [H-3]PAF as a substrate, and proved to be enzymatically active. Lp-PLA(2) inhibitor SB435495 produced a good inhibition curve for inhibition of recombinant Lp-PLA(2) with an IC50 of 57 +/- 1 umol/L. Conclusion: We expressed and purified Lp-PLA(2) at a high level in insect cell-baculovirus expression system. The yield ratio was much greater than that obtained from human plasma and we established a screening model for Lp-PLA(2) inhibitors using the expressed Lp-PLA(2). |
| WOS关键词 | ACTIVATING-FACTOR-ACETYLHYDROLASE ; CORONARY-HEART-DISEASE ; C-REACTIVE PROTEIN ; MIDDLE-AGED MEN ; THERAPEUTIC TARGET ; FOLLOW-UP ; RISK ; INHIBITION ; EVENTS |
| WOS研究方向 | Chemistry ; Pharmacology & Pharmacy |
| 语种 | 英语 |
| CSCD记录号 | CSCD:2462199 |
| WOS记录号 | WOS:000237955200006 |
| 出版者 | ACTA PHARMACOLOGICA SINICA |
| 源URL | [http://119.78.100.183/handle/2S10ELR8/273569]  |
| 专题 | 药理学第一研究室 中科院受体结构与功能重点实验室 新药研究国家重点实验室 |
| 通讯作者 | Wang, Yi-Ping |
| 作者单位 | 1.Chinese Acad Sci, State Key Lab Drug Res, Shanghai Inst Mat Med, Shanghai Inst Biol Sci, Shanghai 201203, Peoples R China 2.Chinese Acad Sci, Dept Med Chem, Shanghai Inst Mat Med, Shanghai Inst Biol Sci, Shanghai 201203, Peoples R China 3.Chinese Acad Sci, Grad Sch, Shanghai 201203, Peoples R China |
| 推荐引用方式 GB/T 7714 | Zhang, Fu-Jun,Cai, Mao-Jun,Shen, Jing-Kang,et al. Expression and purification of lipoprotein-associated phospholipase A(2), a key enzyme involved in atherosclerosis[J]. ACTA PHARMACOLOGICA SINICA,2006,27(6):679-684. |
| APA | Zhang, Fu-Jun,Cai, Mao-Jun,Shen, Jing-Kang,&Wang, Yi-Ping.(2006).Expression and purification of lipoprotein-associated phospholipase A(2), a key enzyme involved in atherosclerosis.ACTA PHARMACOLOGICA SINICA,27(6),679-684. |
| MLA | Zhang, Fu-Jun,et al."Expression and purification of lipoprotein-associated phospholipase A(2), a key enzyme involved in atherosclerosis".ACTA PHARMACOLOGICA SINICA 27.6(2006):679-684. |
入库方式： OAI收割
来源：上海药物研究所
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