Bacterial expression and purification of biologically active human TFF3.
A glutathione S-transferase (GST) fusion protein expression system for the production and purification of recombinant human trefoil factor family-domain peptide 3 (hTFF3) was established. The hTFF3 gene, prepared by PCR, was cloned into a pBluescript KS(+) plasmid, and inserted into a pGEX-4T-1 GST fusion vector. The GST-hTFF3 fusion protein was expressed in Escherichia coli, and hTFF3 was purified with Glutathione Sepharose 4B affinity chromatography, yielding about 3-4 mg of pure hTFF3 in one liter of culture broth. The biological activity of purified hTFF3 was tested in two previously reported rat gastric ulcer models. Oral administration of recombinant hTFF3 has a dose dependent protective effect against ethanol-induced or pylorus ligation-induced gastric mucosa injury in rat, which indicates that our recombinant hTFF3 is biologically active.
Fang
Min
M
Department of Biochemistry and Molecular Biology, National Laboratory of Protein Engineering, College of Life Science, Peking University, Beijing, PR China.
Wang
Wei
W
Wang
Yanru
Y
Ru
Binggen
B
D000818:Animals; D002846:Chromatography, Affinity; D004926:Escherichia coli / Q000378:metabolism; D000431:Ethanol / Q000633:toxicity; D005822:Genetic Vectors / Q000235:genetics*; D005982:Glutathione Transferase / Q000235:genetics; D006801:Humans; D009077:Mucins / Q000302:isolation & purification / Q000494:pharmacology*; D009124:Muscle Proteins / Q000302:isolation & purification / Q000494:pharmacology*; D010455:Peptides; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000494:pharmacology*; D013276:Stomach Ulcer / Q000139:chemically induced / Q000188:drug therapy*; D000071165:Trefoil Factor-3
D009077:Mucins; D009124:Muscle Proteins; D010455:Peptides; D011993:Recombinant Fusion Proteins; C082495:TFF3 protein, human; D000071165:Trefoil Factor-3; D000431:Ethanol; D005982:Glutathione Transferase