High-level production and one-step purification of biologically active human growth hormone in Escherichia coli.
A plasmid has been constructed to direct the synthesis of recombinant human growth hormone (re-hGH) in Escherichia coli as a fusion protein containing a His6 tag at the N-terminus under the control of the T5 promoter. The re-hGH was synthesized in large amounts and accumulated in the form of inclusion bodies upon induction with IPTG. Inclusion bodies were solubilized in 6 M guanidine.HCl and the re-hGH was purified by single-step affinity chromatography on Ni(2+)-nitrilotriacetic acid (NTA) agarose. At the shake flask level, the purified re-hGH was obtained with a yield of 30 mg/l of culture. The re-hGH was biologically active in a node rat lymphoma (Nb2) cell bioassay.
Gene Regulation Laboratory, National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi, India.
R
R
Mukhija
P
P
Rupa
D
D
Pillai
L C
LC
Garg
D000818:Animals; D002846:Chromatography, Affinity; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D013006:Growth Hormone / Q000096:biosynthesis* / Q000302:isolation & purification* / Q000502:physiology; D006639:Histidine; D006801:Humans; D002479:Inclusion Bodies; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D008223:Lymphoma; D008934:Mitogens / Q000502:physiology; D010455:Peptides; D010957:Plasmids / Q000235:genetics; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000302:isolation & purification* / Q000502:physiology; D014407:Tumor Cells, Cultured
D018076:DNA, Complementary; D008934:Mitogens; D010455:Peptides; D011993:Recombinant Fusion Proteins; C033223:polyhistidine; D007544:Isopropyl Thiogalactoside; D006639:Histidine; D013006:Growth Hormone