Expression, purification and characterization of soluble human thrombopoietin receptor from Escherichia coli

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Title
Expression, purification and characterization of soluble human thrombopoietin receptor from Escherichia coli
Author(s)
Heung Rok Park; Ha Na Im; Young Jun Kang; Myeong Hee Yu; Hyo Jeong Hong
Bibliographic Citation
Biotechnology Letters, vol. 22, no. 20, pp. 1611-1617
Publication Year
2000
Abstract
The extracellular domain (edMpl) of human thrombopoietin (TPO) receptor, c-Mpl was expressed in Escherichia coli by changing some nucleotides before and after the translation initiation codon. The mutations increased the expression by approx. 15-fold. The inclusion bodies were solubilized in 8 M guanidine-HCl under reducing conditions and refolded using a glutathione-redox system. The monomeric form of edMpl was purified to near homogeneity by two successive steps of ion-exchange chromatography using DEAE-Sephacel and Mono Q columns. The purified monomeric edMpl inhibited the TPO-dependent cell proliferation, suggesting that it was binding to TPO. Also, antisera raised against the edMpl bound specifically to the soluble receptor secreted by mammalian cells.
Keyword
Inclusion bodiesProtein purificationReceptor domainThrombopoietin
ISSN
0141-5492
Publisher
Springer
DOI
http://dx.doi.org/10.1023/A:1005672824663
Type
Article
Appears in Collections:
1. Journal Articles > Journal Articles
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