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Gene construction, expression, and characterization of double-copy truncated form of human insulin-like growth factor I

   
@article{APS7518,
	author = {He-Ying Sun and Xiao-Hui Liu and Shu-Wen Liang},
	title = {Gene construction, expression, and characterization of double-copy truncated form of human insulin-like growth factor I},
	journal = {Acta Pharmacologica Sinica},
	volume = {22},
	number = {7},
	year = {2016},
	keywords = {},
	abstract = {Aim: To increase the production of recombinant truncated form of insulin-like growth factor I [des(1-3)IGF-I], purify the expressed product, and compare its bioactivity with that of standard IGF-I.
Methods: The second copy of des(1-3)IGF-I gene was inserted into the previously constructed pExSec1/IGF-I to form a pExSec1/2(IGF-I) expression plasmid, then the plasmid was transformed into a protease-deficient E coli strain BL21(DE3). The engineered bacteria were cultured and induced by IPTG at 12 degrees C. The expressed product was purified through ultrafiltration and Sephadex G-50 gelfiltration. The bioactivity of the preliminarily purified protein was tested by MTT method and compared with standard IGF-I.
Results: The amount of des(1-3)IGF-I expressed by pExSec1/2(IGF-I) reached up to 20 % of the total soluble bacterial protein, which was higher than the amount (12 %) expressed by a single copy of pExSec1/IGF-I gene. The purity of recombinant des(1-3)IGF-I reached 49 % and 82 % after ultrafiltration and gelfiltration. The bioactivity of des(1-3)IGF-I after gelfiltration was about 77 % of standard IGF-I at the same concentration.
Conclusion: The yield of recombinant des(1-3)IGF-I was increased about 8 % by construction of expression plasmid with two copies of des(1-3)IGF-I gene compared with only one copy of gene, and preliminarily purified des(1-3)IGF-I showed about 77 % bioactivity compared with standard IGF-I.},
	issn = {1745-7254},	url = {http://www.chinaphar.com/article/view/7518}
}