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Paper Information

Journal:   MEDICAL JOURNAL OF THE ISLAMIC REPUBLIC OF IRAN (MJIRI)   2017 , Volume 31 , Number 1; Page(s) 0 To 0.

Cloning, expression, and spectral analysis of mouse betatrophin



Author(s):  GHOLAMI SAMANEH, Goodarzvand Chegini Koroush, GHEIBI NEMATOLLAH, Mokhtarian Kobra, MOHAMADI MOHSEN, FALAK REZA*
* Immunology Research Center, Iran University of Medical Sciences, Tehran, Iran / Department of Immunology, School of Medicine, Iran University of Medical Sciences, Tehran, Iran
Background: Betatrophin, a novel secretory protein from liver and fatty tissues, is believed to be involved in lipid and glucose metabolism. However, its precise physiological role remains unclear. Here, we report the cloning, expression, and purification steps of mouse betatrophin in a prokaryotic system, followed by its structural analysis. Methods: Specific cloning primers were used to amplify the coding sequence of mouse liver betatrophin. The product was cloned into pET28 and expressed in E. coli BL21 (DE3) cells. The suitability of the refolding procedure was assessed by determining secondary structures of the initial and refolded proteins using circular dichroism spectroscopy. Results: The polymerase chain reaction resulted in a 549 bp nucleotide sequence, encoding a 183 amino acid polypeptide, with an apparent molecular weight of 21 kDa, which was expressed in an inclusion body. Following an optimization and refolding procedure, the recombinant protein was purified by anion exchange and metal affinity chromatography. CD spectra revealed that the refolded protein has suitable configuration. Conclusion: We believe that the produced betatrophin is suitable for further biochemical studies on glucose and lipid metabolism.
Keyword(s): Mouse betatrophin,Recombinant protein,Refolding,CD spectroscopy,Chromatography
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