[Isolation, purification and renaturation of recombinant-DNA-derived porcine somatotropin].
Large scale abstraction and isolation of bacterially synthesized, recombinant-DNA-derived, porcine growth hormone (r-pST) is described. The r-pGH is found in genetic engineering E. coli as the form of inclusion bodies. Pellet fraction which were mainly inclusion bodies, after cell breakage and centrifugation, were collected. Cell envelope components, such as protein, lipid, endotoxin and nucleic acids are selectively removed from the pellet fraction by an EDTA/lysozyme/deoxycholate extraction. Inclusion bodies were dissolved using 6 mol/L guanidine/HCl and air oxidation is then carried out in the presence of the guanidine/HCl. The Guanidine/HCl protein mixture were diluted by renaturation solution. Guanidine/HCl were removed by dialysis and then correctly refolded, oxidized r-pGH were obtained. Injection experiment of hypophysectomized rats proved r-pST with high native bioactivity was obtained.
Li
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Z
Animal Husbandry and Veterinary Research Institute, Shanghai Academy of Agricultural Sciences, Division of Animal Genetic Engineering, Shanghai Municipal Key Laboratory of Agri-Genetics and Breeding, Shanghai 201106, China. zhenli60@public3.sta.net.cn
Yu
R S
RS
Liu
H L
HL
Wang
Y
Y
Zhang
P
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Zhou
Z A
ZA
Zhang
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DF
D000818:Animals; D001681:Biological Assay / Q000379:methods; D004926:Escherichia coli / Q000235:genetics; D013006:Growth Hormone / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D007016:Hypophysectomy / Q000379:methods; D002479:Inclusion Bodies / Q000235:genetics / Q000378:metabolism; D017510:Protein Folding; D020673:Protein Renaturation; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D013552:Swine
D011994:Recombinant Proteins; D013006:Growth Hormone