Large scale preparation of recombinant human parathyroid hormone 1–84 from Escherichia coli

Human parathyroid hormone (hPTH) is a promising agent in the treatment of osteoporosis. The intact recombinant human parathyroid hormone [rhPTH(1–84)] was prepared in a large scale from Escherichia coli using a soluble fusion protein strategy. With degenerate codons, gene of hPTH(1–84) was synthesized, ligated with pET32a(+) vector, and then expressed in E. coli BL21(DE3) cells. The soluble fusion protein His6-thioredoxin-hPTH(1–84) was harvested after purification by immobilized metal affinity chromatography (IMAC). Following enterokinase cleavage, ion-exchange-chromatography (IEC) and size-exclusive-chromatography (SEC) were used, and finally, over 300 mg/l intact hPTH(1–84) with high purity up to 99% was obtained. The purified rhPTH(1–84) was confirmed by mass spectrometry and N-terminal/C-terminal amino-acid sequence analysis. Additionally, this product stimulated adenylate cyclase in Rat Osteosarcoma Cell UMR-106 at the same extent as hPTH standards, indicating that the purified rhPTH(1–84) has full biological activity. The efficient procedure for expression and purification of rhPTH(1–84) may be useful for the mass production of this important protein.