Nematollahi, Leila, Mahboudi, Fereidoun, Azizi, Mohammad, Barkhordari, Farzaneh, Adeli, Ahmad, Khalaj, Vahid (2012). A Comparison of Bone Morphogenetic Protein-7 Mutant Expression in Prokaryotic and Eukaryotic Hosts. , 15(3), 93-102.
Leila Nematollahi; Fereidoun Mahboudi; Mohammad Azizi; Farzaneh Barkhordari; Ahmad Adeli; Vahid Khalaj. "A Comparison of Bone Morphogenetic Protein-7 Mutant Expression in Prokaryotic and Eukaryotic Hosts". , 15, 3, 2012, 93-102.
Nematollahi, Leila, Mahboudi, Fereidoun, Azizi, Mohammad, Barkhordari, Farzaneh, Adeli, Ahmad, Khalaj, Vahid (2012). 'A Comparison of Bone Morphogenetic Protein-7 Mutant Expression in Prokaryotic and Eukaryotic Hosts', , 15(3), pp. 93-102.
Nematollahi, Leila, Mahboudi, Fereidoun, Azizi, Mohammad, Barkhordari, Farzaneh, Adeli, Ahmad, Khalaj, Vahid A Comparison of Bone Morphogenetic Protein-7 Mutant Expression in Prokaryotic and Eukaryotic Hosts. , 2012; 15(3): 93-102.

A Comparison of Bone Morphogenetic Protein-7 Mutant Expression in Prokaryotic and Eukaryotic Hosts

Pathobiology Reserach
Article 8, Volume 15, Issue 3, 2012, Pages 93-102    PDF (1.04 M)
Authors
Leila Nematollahi1; Fereidoun Mahboudi2; Mohammad Azizi3; Farzaneh Barkhordari4; Ahmad Adeli4; Vahid Khalaj* 5
1Ph.D. Candidate, Department of Medical Biotechnology, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran
2Associated Professor, Department of Medical Biotechnology, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran
3Ph.D., Department of Medical Biotechnology, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran
4B.Sc., Department of Medical Biotechnology, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran
5Assistant Professor, Department of Medical Biotechnology, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran
Abstract
Objective: Bone morphogenetic protein-7 (BMP-7) is a multifunctional growth factor predominantly recognized for its osteoinductive properties. Due to the high cost of this protein, the availability of BMP-7 for treatment is limited. The heterologous production of recombinant hBMP-7 has been performed in a number of expression systems. In this study a novel form of BMP-7 was expressed in eukaryotic and prokaryotic hosts. Methods: For expression in the prokaryotic system, the novel protein was secreted to the periplasmic space of Escherichia coli using a pelB signal sequence followed by single-step purification by Ni2+-charged column chromatography. In the mammalian cell expression system, we transferred a full-length cDNA encoding precursor of the novel protein to CHO cells then selected stable clones by using the appropriate antibiotic concentration. Expressions in both systems were confirmed by Western blot analysis. Results: The novel recombinant protein was produced as a 36-38 kDa dimer in the CHO cell line and a 16 kDa monomer in the Escherichia coli system. Quantitative analysis according to ELISA showed that the expression levels of the mutant protein in the eukaryotic and prokaryotic expression systems were 40 ng/ml and 135 ng/ml of the culture media, respectively. Conclusion: In this study, the expression level in Escherichia coli was at least three times more than observed in the CHO cells. However, further optimization is required to obtain a dimer protein in Escherichia coli. The results show that periplasmic expression may be suitable for the production of complex proteins such as BMPs.
Keywords
Escherichia coli; Recombinant Protein; Human bone morphogenetic protein 7; CHO cells
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