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Acta Biochimica et Biophysica Sinica Advance Access originally published online on October 1, 2009
Acta Biochimica et Biophysica Sinica 2009 41(11):900-909; doi:10.1093/abbs/gmp083
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© The Author 2009. Published by ABBS Editorial Office in association with Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.

Development of a soluble PTD-HPV18E7 fusion protein and its functional characterization in eukaryotic cells

Xiaofei Yan1, Shah Walayat1, Qinfeng Shi1, Jin Zheng2 and Yili Wang2,*

1 Center for Cancer Research, the First Affiliated Hospital, Xi'an Jiaotong University, Xi'an 710061, China
2 The Key Laboratory of Biomedical Information Engineering Ministry of Education, Xi'an Jiaotong University, Xi'an 710061, China

* Correspondence address. Tel: +86-29-82655499; Fax: +86-29-82655499; E-mail: wangyili{at}mail.xjtu.edu.cn


  Abstract

Though accumulated evidence has demonstrated the transformation capacity of human papillomavirus (HPV) type 18 protein E7, the underlying mechanism is still arguable. Developing a protein transduction domain (PTD)-linked E7 molecule is a suitable strategy for assessing the biological functions of the protein. In the present study, HPV18 E7 protein fused to an N-terminal PTD was expressed in the form of glutathione S-transferase fusion protein in Escherichia coli with pGEX-4T-3 vector. After glutathione-Sepharose 4B bead affinity purification, immunoblot identification and thrombin cleavage, the PTD-18E7 protein showed structural and functional activity in that it potently transduced the cells and localized into their nuclei. The PTD-18E7 protein transduced the NIH3T3 cells in 30 min and remained stable for at least 24 h. In addition, the PTD-18E7 protein interacted with retinoblastoma protein (pRB) and caused pRB degradation in the transduced NIH3T3 cells. In contrast to the pRB level, p27 protein level was elevated in the transduced NIH3T3 cells. The PTD-18E7 protein gives us a new tool to study the biological functions of the HPV E7 protein.

Keywords    human papillomavirus; E7; fusion protein; transformation

Received: April 22, 2009; Accepted: July 12, 2009
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