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Generation of aminoterminally truncated, stable types of bioactive bovine and porcine fibroblast growth factor 4 in E scherichia coli
Author(s) -
Sugawara Saiko,
Ito Toshihiko,
Sato Shiori,
Sato Yuki,
Sasaki Akira,
Fukuda Tomokazu,
Yamanaka Kenichi,
Sakatani Miki,
Takahashi Masashi,
Kobayashi Masayuki
Publication year - 2014
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1002/bab.1251
Subject(s) - fibroblast growth factor , fibroblast growth factor receptor , biology , phosphorylation , growth factor , escherichia coli , microbiology and biotechnology , signal transduction , kinase , biochemistry , chemistry , receptor , gene
Fibroblast growth factor 4 ( FGF 4) is a crucial growth factor for the development of mammalian embryos. We previously produced hexahistidine‐tagged, bovine and porcine FGF 4 ( P ro 32 to L eu 206 ) proteins without a secretory signal peptide at the aminoterminus in E scherichia coli . Here, we found that these were unstable; site‐specific cleavage between S er 54 and L eu 55 in both FGF 4 derivatives was identified. In order to generate stable FGF 4 derivatives and to investigate their biological activities, aminoterminally truncated and hexahistidine‐tagged bovine and porcine FGF 4 ( L eu 55 to L eu 206 ) proteins, termed H isb FGF 4 L and H isp FGF 4 L , respectively, were produced in E . coli . These FGF 4 derivatives were sufficiently stable and exerted mitogenic activities in fibroblasts. Treatment with the FGF 4 derivatives promoted the phosphorylation of ERK 1/2, which are crucial kinases in the FGF signaling pathway. In the presence of PD 173074, an FGF receptor inhibitor, the phosphorylation of ERK 1/2 was inhibited and resulted in abolition of the growth‐promoting activity of FGF 4 derivatives. Taken together, we demonstrate that H isb FGF 4 L and H isp FGF 4 L are capable of promoting the proliferation of bovine‐ and porcine‐derived cells, respectively, via an authentic FGF signaling pathway. These FGF 4 derivatives may be applicable for dissecting the roles of FGF 4 during embryogenesis in cattle and pigs.