[Cloning, expression and glucose regulation activity of human FGF-21].
Fibroblast growth factor (FGF)-21 is a recently discovered glucose regulator and has potential to become therapeutics for treatment of type 2 diabetes. The aim of this study was to clone and express human FGF-21 gene and characterize its bioactivity for glucose regulation. The hFGF-21 cDNA was cloned from human liver by RT-PCR and subcloned into the pSUMO vector after sequencing confirmation. The recombinant plasmid was transformed into Escherichia coli Rosetta strain. The FGF-21 protein expression was induced by IPTG and purified by Ni-NTA agarose. The FGF-21 product was verified by Western blotting analysis with specific antibody. The bioactivity of the purified protein was examined by glucose uptake assay in 3T3-L1 adipocytes. The cloned hFGF-21 gene consisted of 546 bp, which was in agreement with the published data in GenBank. SDS-PAGE analysis showed that hFGF-21 expressed in the E. coli system was 19.4 kDa in size. The glucose uptake assay in 3T3-L1 adipocytes indicated that the purified hFGF-21 could stimulate glucose uptake in a dose-dependent manner, and glucose transporters (GLUT1) is the functional unit.
Hou
Yu-Ting
YT
Department of Pharmaceutical Biotechnology, College of Life Science, Northeast Agriculture University, Harbin 150030, China.
Li
Jin-Nan
JN
Ren
Gui-Ping
GP
Liu
Ming-Yao
MY
Sun
Guo-Peng
GP
Wang
Wen-Fei
WF
Li
De-Shan
DS
D041721:3T3-L1 Cells; D017667:Adipocytes / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D003001:Cloning, Molecular; D005346:Fibroblast Growth Factors / Q000235:genetics / Q000502:physiology*; D005947:Glucose / Q000378:metabolism*; D051272:Glucose Transporter Type 1 / Q000235:genetics; D051275:Glucose Transporter Type 4 / Q000235:genetics; D051379:Mice; D025842:SUMO-1 Protein / Q000235:genetics
D051272:Glucose Transporter Type 1; D051275:Glucose Transporter Type 4; D025842:SUMO-1 Protein; C414620:fibroblast growth factor 21; D005346:Fibroblast Growth Factors; D005947:Glucose