Recombinant-DNA-derived XXXX growth hormone from Escherichia coli. 1. Demonstration that the hormone is expressed in reduced form, and isolation of the hormone in oxidized, native form.

@entity2097 @entity5828 @entity1520 @entity291 @entity1098
The isolation of bacterially synthesized, recombinant-DNA-derived, @entity291 growth hormone (r-bGH) with native structure is described. The r-bGH is found in insoluble form, in a pellet fraction, after cell breakage and centrifugation. Cell envelope components (protein, lipid, endotoxin) and nucleic acids are selectively removed from the pellet fraction by an @entity1098 /lysozyme/deoxycholate extraction. We demonstrate that the r-bGH is largely reduced until solubilized using 6 M @entity2097 / @entity5828 . Air oxidation is then carried out, in the presence of the @entity2097 / @entity5828 . The oxidation results in a mixture of about one-third @entity1520 -linked oligomers and two-thirds oxidized monomer. The latter may include some incorrectly oxidized material, but appears to be mostly correctly oxidized. The oxidized monomer is isolated by gel filtration in the presence of @entity2097 / @entity5828 . Subsequent @entity2097 / @entity5828 removal leads to refolded, oxidized r-bGH. All steps in the procedure, in particular the oxidation and refolding steps, can be carried out at relatively high protein concentrations.
