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Volume 39, Number 5, October 2006

Secretion of biologically active human epidermal growth factor from Escherichia coli using Yersinia pestis Caf1 signal peptide


Yi-Lin Liu, Liang-Min Huang, Wen-Po Lin, Chung-Chin Tsai, Tsung-Shun Lin, Yu-Hua Hu, Hquan-Shin Chen, Jun-Ming Han, Hsian-Jenn Wang, Yu-Tien Liu
Department of Dermatology, Taipei Veterans General Hospital, Taipei; and Institute of Microbiology and Immunology, National Defense Medical Center, Taipei, Taiwan

Received: July 9, 2005    Revised: January 14, 2006    Accepted: February 6, 2006   

 

Corresponding author:

Yu-Tien Liu, Ph.D., Professor of Microbiology, Institute of Microbiology and Immunology, National Defense Medical Center, P.O. Box 90048-505, Neihu, Taipei, Taiwan. E-mail: ytliu@mail.ndmctsgh.edu.tw This e-mail address is being protected from spam bots, you need JavaScript enabled to view it

 



 

Background and purpose: 

The Caf1 secretion pathway of Yersinia pestis is one of the most well-characterized export machineries. To facilitate the secretion of human epidermal growth factor (hEGF) in Escherichia coli, a DNA fragment containing the synthetic gene for hEGF was joined to a sequence encoding the signal peptide of Yersinia pestis Caf1 protein.

 



 

Methods:

The gene for hEGF was synthesized by overlapping polymerase chain reaction technique and was placed under the control of the caf1 gene promoter in the recombinant plasmid pHL401 which was used to transfect E. coli BL-21 for production of hEGF. The biological function of recombinant hEGF was measured by estimating its ability to stimulate the proliferation of human embryonic kidney-293 cells.

 



 

Results:

The results indicated that the expressed hybrid protein was processed during the secretion process. The majority of the mature hEGF was recovered from the periplasm and medium fractions, with a small amount of the expressed hEGF deposited in the cytoplasm. Furthermore, it was found that the cell proliferation was enhanced by the recombinant hEGF.

 



 

Conclusion:

These results suggested that the recombinant hEGF was successfully secreted through the inner membrane of cells into the periplasm and then through the outer membrane into the medium via the action of the signal peptide of Y. pestis Caf1 in E. coli. The mitogenic activity of hEGF in cells was demonstrated.

 



 

Key words:

Bacterial proteins, Caf1, protein sorting signals, recombinant fusion proteins, Yersinia pestis



 

J Microbiol Immunol Infect 2006;39:366-371.