[Prokaryotic expression and purification of human hepatic stimulator substance].
To explore the possibility of prokaryotic expression of human hepatic stimulator substance (hHSS), hHSS gene was inserted in the downstream of glutathion S-transferase (GST) in a pET-42a expression vector and recombinant GST-hHSS fusion protein was expressed under IPTG induction in BL-21(DE3) cells. The recombinant HSS was purified with His.Tag affinity chromatography, and its bioactivity was analyzed. The results showed that GST-hHSS fusion protein was expressed both as a soluble or a inclusive body in bacterial cytosol. The soluble GST-hHSS expression reached up to 30% of the whole soluble protein of bacteria as determined by densitometry. The cleavage of GST-hHSS fusion protein with Factor Xa produced two fragments of the protein, which sized 33 and 15 kD, respectively. The molecular weight of recombinant HSS protein was identical to theoretical deduction based on the DNA sequences. The protein homology of 15 kD hHSS could be efficiently eluted out after Factor Xa cleavage. It is further indicated that the recombinant hHSS is able to proliferate hepatoma cells of BEL-7402 in the preliminary experiments.
Du
Hai-Jun
HJ
Department of Cell Biology, Capital University of Medical Sciences, Beijing 100054, anwei@cpums.edu.cn
Sun
Hong-Liu
HL
Chen
Li
L
An
Wei
W
D002455:Cell Division / Q000187:drug effects; D015870:Gene Expression; D005982:Glutathione Transferase / Q000235:genetics; D006133:Growth Substances / Q000235:genetics* / Q000302:isolation & purification* / Q000494:pharmacology; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D010455:Peptides / Q000235:genetics* / Q000302:isolation & purification* / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D006133:Growth Substances; D036341:Intercellular Signaling Peptides and Proteins; D010455:Peptides; D011993:Recombinant Fusion Proteins; C073456:hepatic stimulator substance; D005982:Glutathione Transferase