Optimization of Prokaryotic Expression Conditions of Human β2-microglobulin in <i>E. Coli</i> and Its Purification_Journal of Biomedical Engineering

Authors：

JIAOLiyuan , CAILei , RENYanna , ZHAOXiaoni ,  WANGJihua

Guangzhou Wondfo Biotech Co. Ltd., Guangzhou 510663, China;

Corresponding author：

WANGJihua, Email: jhwang@wondfo.com.cn

Keywords：

β2-microglobulin; prokaryotic expression; optimization; purification; Western blot

DOI：

10.7507/1001-5515.20150186

Video：

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To obtain recombinant human β2-microglobulin (rhβ2M) with properties of good solubility and high purity from E.coli, prokaryotic expression conditions were optimized and protein purification was performed in this study. After testing the effect of different IPTG concentrations, temperatures and induction times on the production of rhβ2M, the optimum expression conditions were determined, i.e. joining IPTG to final concentration being 0.8 mmol/L and inducing time 6 h and at temperature of 25℃. Under the optimum induction conditions, the ratio of soluble rhβ2M to soluble bacterial protein was 63.7%. After purified by Ni Sepharose 6 Fast Flow, the purity of rhβ2M achieved a greater value of 95%. Western blot analysis revealed that rhβ2M possessed the antigen property that specifically interacted with anti-β2M antibody.

Citation： JIAOLiyuan, CAILei, RENYanna, ZHAOXiaoni, WANGJihua. Optimization of Prokaryotic Expression Conditions of Human β2-microglobulin in E. Coli and Its Purification. Journal of Biomedical Engineering, 2015, 32(5): 1050-1055. doi: 10.7507/1001-5515.20150186 Copy

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9.	何贤辉,徐丽慧,刘毅,等.人β2微球蛋白基因克隆及其在大肠杆菌中的高效表达[J].生物工程学报,2004,20(1):99-103.
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11.	张长庚,姚新杰,王文智,等.人β2微球蛋白在pET32a表达载体中的克隆与表达[J].国际检验医学杂志,2010,31(12):1390-1391.
12.	朴文花,张恒辉,何豫,等.人β2微球蛋白在pET-22b(+)表达载体中的克隆、表达与纯化[J].中国免疫学杂志,2005,21(2):99-101.
13.	APPAY V, ROWLANF-JONES S L. The assessment of antigen-specific CD8+ T cells through the combination of MHC class I tetramer and intracellular staining[J]. J Immunol Methods, 2002, 268(1):9-19.
14.	商庆龙,马延秀,郭志伟,等.正交试验在人乳头瘤病毒16型E2蛋白原核表达优化中的应用[J].生物医学工程学杂志,2011,28(5):988-991.
15.	黄演婷,卢雪梅,汪洁,等.肝靶向干扰素(IFN-CSP)融合基因原核表达条件的优化[J].生物医学工程学杂志,2014,31(2):432-438.

1. SHIELDS M J, KUBOTA R, HODGSON W, et al. The effect of human β2-microglobulin on major histocompatibility complex I peptide loading and the engineering of a high affinity variant[J]. J Biol Chem, 1998, 273(43):28010-28018.
2. LI K, DU H F, LIAN X W, et al. Characterization of β2-microglobulin expression in different types of breast cancer[J]. BMC Cancer, 2014, 14:750-758.
3. ASTOR B C, MUTH B, KAUFMAN D B, et al. Serum β2-microglobulin at discharge predicts mortality and graft loss following kidkey transplantation[J]. Kidney international, 2013, 84(4):810-817.
4. NGUYEN C, DREX S, HEIDET L, et al. Fetal serum α-1microglobulin for renal function assessment:comparison with β2-microglobulin and cystatun C[J]. Prenatal Diagnosis,2013, 33(8):775-781.
5. TSIMBERDOU A M, KANTARJIAN H M, WEN S, et al. The prognostic significance of serum β2-microglobulin levels in acute myloid leukemia and prognostic scores predicting survival:analysis if 1180 patients[J]. Clin Cancer Res, 2008, 14(3):721-730.
6. JOSSON S, MATSUOKA Y, GURURAJAN M, et al. Inhibition of β2-microglobulin/Hemochromatosis enhances radiation sensitivity by induction of iron overload in prostate cancer cells[J]. PLOS ONE, 2013, 8(7):e68366.
7. NEAD K T, MPHIL, ZHOU M J, et al. Usefulness of the addition of beta-2-microglobulin, cystatin C and C-Reactive protein on an established risk factors model to improve mortality risk prediction in patients undergoing coronary angiography[J]. The American Journal of Cardiology, 2013, 111(6):851-856.
8. HUANG W C, ZHOU H E, CHUNG L W, et al. Androgen receptor survival signaling is blocked by anti-β2-microglobulin monoclonal antibody via a mitogen-activated protein kinase/lipogenic pathway in human prostate cancer cells[J]. J Biol Chem, 2010, 285(11):7947-7956.
9. 何贤辉,徐丽慧,刘毅,等.人β2微球蛋白基因克隆及其在大肠杆菌中的高效表达[J].生物工程学报,2004,20(1):99-103.
10. 张劲,郭蔼光,丰美福,等.人β2微球蛋白在大肠杆菌中的表达与优[J].西北农林科技大学学报(自然科学版),2006,34(7):87-90.
11. 张长庚,姚新杰,王文智,等.人β2微球蛋白在pET32a表达载体中的克隆与表达[J].国际检验医学杂志,2010,31(12):1390-1391.
12. 朴文花,张恒辉,何豫,等.人β2微球蛋白在pET-22b(+)表达载体中的克隆、表达与纯化[J].中国免疫学杂志,2005,21(2):99-101.
13. APPAY V, ROWLANF-JONES S L. The assessment of antigen-specific CD8+ T cells through the combination of MHC class I tetramer and intracellular staining[J]. J Immunol Methods, 2002, 268(1):9-19.
14. 商庆龙,马延秀,郭志伟,等.正交试验在人乳头瘤病毒16型E2蛋白原核表达优化中的应用[J].生物医学工程学杂志,2011,28(5):988-991.
15. 黄演婷,卢雪梅,汪洁,等.肝靶向干扰素(IFN-CSP)融合基因原核表达条件的优化[J].生物医学工程学杂志,2014,31(2):432-438.

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