Мақаланы E-mail арқылы жіберу Recombinant human erythropoietin with additional processable protein domains: Purification of protein synthesized in Escherichia coli heterologous expression system
- Авторлар: Grunina T.1, Demidenko A.1, Lyaschuk A.1, Poponova M.1, Galushkina Z.1, Soboleva L.1, Cherepushkin S.2, Polyakov N.1,3, Grumov D.1, Solovyev A.1, Zhukhovitsky V.1,4, Boksha I.1,5, Subbotina M.1,6, Gromov A.1, Lunin V.1,6, Karyagina A.1,6,7
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Мекемелер:
- Gamaleya National Research Center of Epidemiology and Microbiology
- State Research Institute of Genetics and Selection of Industrial Microorganisms
- Vernadsky Institute of Geochemistry and Analytical Chemistry
- Sechenov Moscow State Medical University
- Mental Health Research Center
- All-Russia Research Institute of Agricultural Biotechnology
- Belozersky Institute of Physico-Chemical Biology
- Шығарылым: Том 82, № 11 (2017)
- Беттер: 1285-1294
- Бөлім: Article
- URL: https://journals.rcsi.science/0006-2979/article/view/151503
- DOI: https://doi.org/10.1134/S0006297917110062
- ID: 151503
Дәйексөз келтіру
Ашық рұқсат
Рұқсат берілді
Тек жазылушылар үшін
Аннотация
Three variants of human recombinant erythropoietin (rhEPO) with additional N-terminal protein domains were obtained by synthesis in an Escherichia coli heterologous expression system. These domains included (i) maltose-binding protein (MBP), (ii) MBP with six histidine residues (6His) in N-terminal position, (iii) s-tag (15-a.a. oligopeptide derived from bovine pancreatic ribonuclease A) with N-terminal 6His. Both variants of the chimeric protein containing MBP domain were prone to aggregation under nondenaturing conditions, and further purification of EPO after the domain cleavage by enterokinase proved to be impossible. In the case of 6His-s-tag-EPO chimeric protein, the products obtained after cleavage with enterokinase were successfully separated by column chromatography, and rhEPO without additional domains was obtained. Results of MALDI-TOF mass spectrometry showed that after refolding 6His-s-tag-EPO formed a structure similar to that of one of native EPO with two disulfide bonds. Both 6His-s-tag-EPO and rhEPO without additional protein domains purified after proteolysis possessed the same biological activity in vitro in the cell culture.
Негізгі сөздер
Авторлар туралы
T. Grunina
Gamaleya National Research Center of Epidemiology and Microbiology
Email: akaryagina@gmail.com
Ресей, Moscow, 123098
A. Demidenko
Gamaleya National Research Center of Epidemiology and Microbiology
Email: akaryagina@gmail.com
Ресей, Moscow, 123098
A. Lyaschuk
Gamaleya National Research Center of Epidemiology and Microbiology
Email: akaryagina@gmail.com
Ресей, Moscow, 123098
M. Poponova
Gamaleya National Research Center of Epidemiology and Microbiology
Email: akaryagina@gmail.com
Ресей, Moscow, 123098
Z. Galushkina
Gamaleya National Research Center of Epidemiology and Microbiology
Email: akaryagina@gmail.com
Ресей, Moscow, 123098
L. Soboleva
Gamaleya National Research Center of Epidemiology and Microbiology
Email: akaryagina@gmail.com
Ресей, Moscow, 123098
S. Cherepushkin
State Research Institute of Genetics and Selection of Industrial Microorganisms
Email: akaryagina@gmail.com
Ресей, Moscow, 117545
N. Polyakov
Gamaleya National Research Center of Epidemiology and Microbiology; Vernadsky Institute of Geochemistry and Analytical Chemistry
Email: akaryagina@gmail.com
Ресей, Moscow, 123098; Moscow, 119991
D. Grumov
Gamaleya National Research Center of Epidemiology and Microbiology
Email: akaryagina@gmail.com
Ресей, Moscow, 123098
A. Solovyev
Gamaleya National Research Center of Epidemiology and Microbiology
Email: akaryagina@gmail.com
Ресей, Moscow, 123098
V. Zhukhovitsky
Gamaleya National Research Center of Epidemiology and Microbiology; Sechenov Moscow State Medical University
Email: akaryagina@gmail.com
Ресей, Moscow, 123098; Moscow, 119991
I. Boksha
Gamaleya National Research Center of Epidemiology and Microbiology; Mental Health Research Center
Email: akaryagina@gmail.com
Ресей, Moscow, 123098; Moscow, 115522
M. Subbotina
Gamaleya National Research Center of Epidemiology and Microbiology; All-Russia Research Institute of Agricultural Biotechnology
Email: akaryagina@gmail.com
Ресей, Moscow, 123098; Moscow, 127550
A. Gromov
Gamaleya National Research Center of Epidemiology and Microbiology
Хат алмасуға жауапты Автор.
Email: alexander.v.gromov@gmail.com
Ресей, Moscow, 123098
V. Lunin
Gamaleya National Research Center of Epidemiology and Microbiology; All-Russia Research Institute of Agricultural Biotechnology
Email: akaryagina@gmail.com
Ресей, Moscow, 123098; Moscow, 127550
A. Karyagina
Gamaleya National Research Center of Epidemiology and Microbiology; All-Russia Research Institute of Agricultural Biotechnology; Belozersky Institute of Physico-Chemical Biology
Хат алмасуға жауапты Автор.
Email: akaryagina@gmail.com
Ресей, Moscow, 123098; Moscow, 127550; Moscow, 119992
