Expression and Purification of Bioactive High-Purity Recombinant Mouse SPP1 in Escherichia coli
Expression and Purification of Bioactive High-Purity Recombinant Mouse SPP1 in Escherichia coli
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具有生物活性的高纯度重组小鼠SPP1在大肠杆菌中的表达和纯化
DOI：
10.1007/s12010-014-0849-7
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发表时间：
2011-03
影响因子：
3
通讯作者：
Yu, Yan
中科院分区：
文献类型：
--
作者：
Gao, Jin;Li, Jingjing;Han, Wei;Yu, Yan
分泌型磷蛋白1（SPP 1）是一种磷酸化的酸性糖蛋白。它在多种组织中广泛表达，并且参与许多生理和病理事件，包括癌症转移、组织重塑、促炎症调节和细胞存活。SPP 1具有保护组织和器官免受损伤和创伤的功能，使其本身有可能成为治疗靶点，或使其抗体或其他对抗试剂有可能成为候选药物。非标记的（天然的）重组SPP 1在治疗和药物研究中将是有价值的。本研究将不含信号肽的小鼠Spp 1 DNA片段构建到pET 28 a（+）载体中，转化大肠杆菌BL 21（DE 3）。重组小鼠SPP 1（rmSPP 1）在异丙基β-d-硫代半乳糖苷（IPTG）诱导下在细菌中表达。采用等电沉淀法和硫酸铵分级分离法提高rmSPP 1的丰度，并采用阴离子和阳离子交换色谱法进一步纯化rmSPP 1。最后得到了产率为12.8%、纯度为97%、具有良好生物活性、内毒素含量低的rmSPP 1。
Secreted phosphoprotein 1 (SPP1) is a phosphorylated acidic glycoprotein. It is broadly expressed in a variety of tissues, and it is involved in a number of physiological and pathological events, including cancer metastasis, tissues remodeling, pro-inflammation regulation, and cell survival. SPP1 has shown its function of protecting tissues and organs against injury and wound, giving itself potentials to become a therapy target or giving its antibodies of other counter-acting reagents potentials to become drug candidates. Non-tagged (native) recombinant SPP1 would be valuable in therapeutic and pharmaceutical researches. In our study, mouse Spp1 DNA fragment without signal peptide was built in pET28a(+) vector and transformed into Escherichia coli BL21 (DE3). The recombinant mouse SPP1 (rmSPP1) was then expressed in bacteria upon induction by isopropyl β-d-thiogalactopyranoside (IPTG). The abundance of rmSPP1 was increased using isoelectric precipitation and ammonium sulfate fractionation methods, and anion and cation exchange chromatography was employed to further purify rmSPP1. Finally, we got rmSPP1 product with 12.8 % productivity, 97 % purity, satisfactory bioactivity, and low endotoxin content.
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影响因子：
11.2
作者：
Pelicano H;Lu W;Zhou Y;Zhang W;Chen Z;Hu Y;Huang P
通讯作者：
Huang P
影响因子：
3.9
作者：
Kaori Sato;S. Ozawa;K. Izukuri;Y. Kato;R. Hata
通讯作者：
Kaori Sato;S. Ozawa;K. Izukuri;Y. Kato;R. Hata
影响因子：
1.6
作者：
S. Sørensen;S. J. Justesen;A. Johnsen
通讯作者：
S. Sørensen;S. J. Justesen;A. Johnsen
DOI：
10.1042/bj2690679
发表时间：
1990-08
期刊：
The Biochemical journal
影响因子：
--
作者：
Z. Abraham;N. Symonds
通讯作者：
Z. Abraham;N. Symonds
影响因子：
4.8
作者：
Nara, Noriko;Nakayama, Yuki;Hara, Takahiko
通讯作者：
Hara, Takahiko