ISSN 2410-7751 (Print)
ISSN 2410-776X (Online)

"Biotechnologia Acta" V. 8, No 4, 2015
Р. 98-107, Bibliography 19, Ukrainian
Universal Decimal Classification: 577(112+25+354.9)+57(085.2+086.2)
https://doi.org/10.15407/biotech8.04.098

 OBTAINING OF RECOMBINANT ANALOGUE OF HUMAN SHB-EGF USING OF TRANSPOSON-TRANSPOSASE GENETIC SYSTEM

 N. V. Korotkevych, A. Yu. Labyntsev, D. V. Kolybo, S. V. Komisarenko

Palladin Institute of Biochemistry of the National Academy of Scienses of Ukraine, Kyiv

The goal of work was obtaining of recombinant soluble form of heparinbinding EGF-like growth factor sHB-EGF in eukaryotic cells based on transposon genetic system. We used methods of recombinant DNA and obtaining of eukaryotic cells for  producing of recombinant proteins. We have obtained stable eukaryotic producer of recombinant analogue of soluble form of human heparinbinding EGF-like growth factor sHB-EGF based on HEK293 cells. We used transposon based non-viral expression system for sHBEGF expression. Moreover, studies in mice have shown the possibility of using sHB-EGF expressing genetic constructs for in vivo gene therapy of various pathological conditions.

Key words: heparin-binding EGF-like growth factor HB-EGF; transposase; recombinant proteins; growth factors.

© Palladin Institute of Biochemistry of the National Academy of Sciences of Ukraine, 2015

{accordion title="References" open="false"}

1. Higashiyama S., Abraham J. A., Miller J., Fiddes J. C., Klagsbrun M. A heparin-binding growth factor secreted by macrophage-like cells that is related to EGF. Science. 1991, 251(251), 936–939. doi:10.1126/science.1840698

2. Raab G., Klagsbrun M. Heparin-binding EGFlike growth factor. Biochimica et biophysica acta. 1997, 1333(3): F179–199. doi:10.1016/S0304-419X(97)00024-3

3. Higashiyama S., Nanba D. ADAM-mediated ectodomain shedding of HB-EGF in receptor cross-talk. Biochim Biophys Acta. 2005, 1751(1), 110-117. doi:10.1016/j.bbapap.2004.11.009

4. Hieda M., Isokane M., Koizumi M., Higashi C., Tachibana T., Shudou M., Taguchi T., Hieda Y., Higashiyama S. Membrane-anchored growth factor, HB-EGF, on the cell surface targeted to the inner nuclear membrane. The Journal of Cell Biology. 2008, 180(4), 763–769. doi: 10.1083/jcb.200710022

5. Higashiyama S., Iwabuki H., Morimoto C., Hieda M., Inoue H., Matsushita N. Membraneanchored growth factors, the epidermal growth factor family: beyond receptor ligands. Cancer Science. 2008, 99(2), 214–220.
doi: 10.1111/j.1349-7006.2007.00676.x.

6. Harris R. C., Chung E., Coffey R. J. EGFreceptor ligands. Experimental cell research. 2003, 284(1), 2–13. doi:10.1016/S0014-4827(02)00105-2

7. Naglich J., Metherall E., Russell D., Eidels L. Expression cloning of a diphtheria toxin receptor: identity with a heparin-binding EGF-like growth factor precursor. Cell. 1992, 69(6), 1051–1061. doi:10.1016/0092-8674(92)90623-K

8. Shirakata Y., Kimura R., Nanba D., Iwamoto R., Tokumaru S., Morimoto C., Yokota K., Nakamura M., Sayama K., Mekada E., Higashiyama S., Hashimoto K. Heparin-binding EGF-like growth factor accelerates keratinocyte migration and skin wound healing. Journal of Cell Science. 2005, 118(11), 2363–2370. doi: 10.1242/ jcs.02346

9. Singh B., Tsukada T., Zent R., Harris C. Membrane-associated HB-EGF modulates HGF-induced cellular responses in MDCK cells. Journal of Cell Science. 2004, 117(8), 1365–1379. doi: 10.1242/ jcs.01037

10. Zarrouki B., Benterki I., Font?s G., Peyot M., Seda O., Prentki M., Poitout V. Epidermal growth factor receptor signaling promotes pancreatic ?-cell proliferation in response to nutrient excess in rats through mTOR and FOXM1. Diabetes. 2014, 63(3), 982–993. doi: 10.2337/db13-0425

11. Kozawa J., Tokui Y., Moriwaki M., Li M., Ohmoto H., Yuan M., Zhang J., Iwahashi H., Imagawa A., Yamagata K., Tochino Y., Shimomura I., Higashiyama S., Miyagawa J. Regenerative and therapeutic effects of heparin-binding epidermal growth factor-like growth factor on diabetes by gene transduction through retrograde pancreatic duct injection of adenovirus vector. Pancreas. 2005, 31(1), 32–42.
doi: 10.1097/01.mpa.0000163177.59920.f8

12. Kiso S., Kawata S., Tamura S., Inui Y., Yoshida Y., Sawai Y., Umeki S., Ito N., Yamada A., Miyagawa J., Higashiyama S., Iwawaki T., Saito M., Taniguchi N., Matsuzawa Y., Kohno K. Liver regeneration in heparin-binding EGF-like growth factor transgenic mice after partial hepatectomy. Gastroenterology. 2003, 124(3), 701–707. doi:10.1053/gast.2003.50097

13. Korotkevych N. V., Kolybo D. V., Labyntsev А. Yu., Romaniuk S. І., Komisarenko S. V. Obtaining of recombinant human heparin-binding EGFlike growth factor and perspectives of its application in biotechnology. Biotekhnolohiia. 2010, 3(4), 44–54. (In Ukrainian).

14. Aronovich E., McIvor R., Hackett P. The Sleeping Beauty transposon system: a nonviral vector for gene therapy. Human Molecular Genetics. 2011, 20(R1), R14–R20. doi: 10.1093/hmg/ddr140

15. Bell J., Podetz-Pedersen K., Aronovich E., Belur L., McIvor S., Hackett P. Preferential delivery of the Sleeping Beauty transposon system to livers of mice by hydrodynamic injection. Nature protocols. 2007, 2(12), 3153–3165. doi: 10.1038/nprot.2007.471

16. Green M., Sambrook J. Molecular cloning: a laboratory manual. N.-Y.: Cold Spring Harbor Laboratory. 2012, 726 p.

17. Dower J., Miller F., Ragsdale W. High efficiency transformation of E. coli by high voltage electroporation. Nucleic Acids Research. 1988, 16(13), 6127–6145. http://dx.doi.org/10.1093/nar/16.13.6127

18. Harlow E., Lane D. Antibodies: a laboratory manual. N.-Y.: Cold Spring Harbor Laboratory. 1988, 720 p.

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