The Optimization of Soluble PTEN Expression in Escherichia coli



Yamei Hu1, Yang An1, Na Fang1, Yanzhang Li1, Haiying Jin2, Adil Nazarali3, *, Shaoping Ji1, *
1 Department of Biochemistry and Molecular Biology, Medical School, Henan University, China
2 Department of Interventional Therapy, Huaihe Hospital, Henan University, China
3 College of Pharmacy and Nutrition and Neuroscience Research Cluster, University of Saskatchewan, Canada


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© Hu et al.; Licensee Bentham Open.

open-access license: This is an open access article licensed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted, non-commercial use, distribution and reproduction in any medium, provided the work is properly cited.

* Address correspondence to these authors at the Department of Biochemistry and Molecular Biology, Medical School, Henan University, China; Tel: 86-371-23888286; Fax: 86-371-23880585; E-mails: aj.nazarali@usask.ca;jishaoping@hotmail.com


Abstract

As a vital tumor suppressor, PTEN (Phosphatase and tension homolog deleted on chromosome 10) is involved in inherited syndromes, and is among the most frequently inactivated tumor suppressor gene in sporadic cancers. PTEN loss-of-function widely occurs in human cancers via a variety of mechanisms, including genetic alterations and posttranslational modification. These suggest PTEN has a role of functional importance in a variety of cancers. In the present study, we constructed a prokaryotic expression vector that efficiently expresses GST-PTEN (the target protein in which PTEN is fused with glutathione S-transferase tag) in E. coli. We found that the target protein was partially soluble although major portions of the protein remained in the inclusion bodies. Furthermore, we explored the optimal induction temperature, isopropyl β-D-1-thiogalactopyranoside (IPTG) concentration and induction time in a series of experiments. Expression level analysis indicated that PTEN reached its peak level at 36C for 8 h with 1.5625mM IPTG, while solubility analysis revealed the optimal induction temperature was at 20C, the optimal IPTG concentration was 0.1µM and the optimal induction time was up to 8 h. Taken together, we provide an optimal induction condition for expressing soluble fusion protein of PTEN in E. coli, facilitating further analysis of PTEN’s biological function in vitro.

Keywords: Expression, GST-PTEN, purification, solubility analysis.