Enviar artigo por via de e-mail Optimization of rhBMP-2 active-form production in a heterologous expression system using microbiological and molecular genetic approaches
- Autores: Karyagina A.S.1,2,3, Boksha I.S.1, Grunina T.M.1, Demidenko A.V.1, Poponova M.S.1, Sergienko O.V.1,3, Lyaschuk A.M.1, Galushkina Z.M.1, Soboleva L.A.1, Osidak E.O.1,4, Semikhin A.S.1, Gromov A.V.1, Lunin V.G.1,3
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Afiliações:
- Gamaleya Federal Research Center for Epidemiology and Microbiology
- Belozersky Research Institute of Physico-Chemical Biology
- All-Russia Research Institute of Agricultural Biotechnology
- Imtek Ltd.
- Edição: Volume 31, Nº 4 (2016)
- Páginas: 208-213
- Seção: Experimental Works
- URL: https://journals.rcsi.science/0891-4168/article/view/178130
- DOI: https://doi.org/10.3103/S0891416816040030
- ID: 178130
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Resumo
Recombinant bone morphogenetic protein-2 (rhBMP-2) has pronounced osteoinductive properties, as evidenced by the results of experimental and clinical practices. This applies to both the protein produced in eukaryotic cells and the protein synthesized in bacterial cells. In eukaryotic expression systems, production of the protein is extremely low and, consequently, the cost of materials on its basis is very high. Therefore, optimization of heterologous expression systems for rhBMP-2 production represents an important task. In the present work, optimization of codon composition of the rhBMP-2 gene nucleotide sequence and secondary structure of the transcript, as well as strain selection for efficient gene expression, were carried out. The producing strain based on Escherichia coli BL-21(DE3) provides a high level of rhBMP-2 synthesis (about 57% of total cell proteins). Biological activity of rhBMP-2 dimeric forms purified from the obtained producing strain was measured by induction of alkaline phosphatase activity in C2C12 cells. It is comparable with that of commercial rhBMP-2 expressed in E. coli (R&D Systems, United States). Purified rhBMP-2 does not contain impurities of E. coli endotoxin and can be used in experimental studies of osteoinduction in laboratory animals.
Sobre autores
A. Karyagina
Gamaleya Federal Research Center for Epidemiology and Microbiology; Belozersky Research Institute of Physico-Chemical Biology; All-Russia Research Institute of Agricultural Biotechnology
Autor responsável pela correspondência
Email: akaryagina@gmail.com
Rússia, Moscow, 123098; Moscow, 119991; Moscow, 127550
I. Boksha
Gamaleya Federal Research Center for Epidemiology and Microbiology
Email: akaryagina@gmail.com
Rússia, Moscow, 123098
T. Grunina
Gamaleya Federal Research Center for Epidemiology and Microbiology
Email: akaryagina@gmail.com
Rússia, Moscow, 123098
A. Demidenko
Gamaleya Federal Research Center for Epidemiology and Microbiology
Email: akaryagina@gmail.com
Rússia, Moscow, 123098
M. Poponova
Gamaleya Federal Research Center for Epidemiology and Microbiology
Email: akaryagina@gmail.com
Rússia, Moscow, 123098
O. Sergienko
Gamaleya Federal Research Center for Epidemiology and Microbiology; All-Russia Research Institute of Agricultural Biotechnology
Email: akaryagina@gmail.com
Rússia, Moscow, 123098; Moscow, 127550
A. Lyaschuk
Gamaleya Federal Research Center for Epidemiology and Microbiology
Email: akaryagina@gmail.com
Rússia, Moscow, 123098
Z. Galushkina
Gamaleya Federal Research Center for Epidemiology and Microbiology
Email: akaryagina@gmail.com
Rússia, Moscow, 123098
L. Soboleva
Gamaleya Federal Research Center for Epidemiology and Microbiology
Email: akaryagina@gmail.com
Rússia, Moscow, 123098
E. Osidak
Gamaleya Federal Research Center for Epidemiology and Microbiology; Imtek Ltd.
Email: akaryagina@gmail.com
Rússia, Moscow, 123098; Moscow, 121552
A. Semikhin
Gamaleya Federal Research Center for Epidemiology and Microbiology
Email: akaryagina@gmail.com
Rússia, Moscow, 123098
A. Gromov
Gamaleya Federal Research Center for Epidemiology and Microbiology
Email: akaryagina@gmail.com
Rússia, Moscow, 123098
V. Lunin
Gamaleya Federal Research Center for Epidemiology and Microbiology; All-Russia Research Institute of Agricultural Biotechnology
Email: akaryagina@gmail.com
Rússia, Moscow, 123098; Moscow, 127550
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