Human parathyroid hormone (hPTH) shows great potential as a therapeutic option for osteoporosis in patients who do not respond sufficiently to conventional therapies. We utilize recombinant DNA technology to enhance the drug development process and introduce a state-of-the-art purification protocol for the first 34 residues of human parathyroid hormone (hPTH), as these residues are responsible for the majority of its hormonal activity. Typically, the purification of recombinant human parathyroid hormone has relied on His-Tag fusion partners using Ni²⁺-chelating chromatography. This is the first time ion exchange chromatography has been used for the soluble PTH form, and no reports of ion exchange chromatography for this purpose have been documented. SUMO-tag technology and a three-step chromatography process were used to purify recombinant human parathyroid hormone [rhPTH<CitationRef AdditionalCitationIDS="CR2 CR3 CR4 CR5 CR6 CR7 CR8 CR9 CR10 CR11 CR12 CR13 CR14 CR15 CR16 CR17 CR18 CR19 CR20 CR21 CR22 CR23 CR24 CR25 CR26 CR27 CR28 CR29 CR30 CR31 CR32 CR33" CitationID="CR1">1</CitationRef>–<CitationRef CitationID="CR34">34</CitationRef> on a large scale from Escherichia coli. The rhPTH (1–84) construct was cloned into E. coli host cells and expressed at fermenter scale with a 5-liter volume. The SUMO-hPTH<CitationRef AdditionalCitationIDS="CR2 CR3 CR4 CR5 CR6 CR7 CR8 CR9 CR10 CR11 CR12 CR13 CR14 CR15 CR16 CR17 CR18 CR19 CR20 CR21 CR22 CR23 CR24 CR25 CR26 CR27 CR28 CR29 CR30 CR31 CR32 CR33" CitationID="CR1">1</CitationRef>–<CitationRef CitationID="CR34">34</CitationRef> fusion protein was extracted and purified using a Capto™ adhere column. The N-terminus of SUMO-hPTH was specifically cleaved with SUMO protease, after which rhPTH was further purified through sequential chromatography on Capto adhere and Capto SP columns. Our purification procedure for rhPTH from E. coli achieved over 98% purity, as confirmed by SDS-PAGE and RP-HPLC. This study demonstrates that the purified rhPTH<CitationRef AdditionalCitationIDS="CR2 CR3 CR4 CR5 CR6 CR7 CR8 CR9 CR10 CR11 CR12 CR13 CR14 CR15 CR16 CR17 CR18 CR19 CR20 CR21 CR22 CR23 CR24 CR25 CR26 CR27 CR28 CR29 CR30 CR31 CR32 CR33" CitationID="CR1">1</CitationRef>–<CitationRef CitationID="CR34">34</CitationRef> meets the purity standards for commercial drug specifications. We propose that our three-step chromatography process effectively purifies active rhPTH<CitationRef AdditionalCitationIDS="CR2 CR3 CR4 CR5 CR6 CR7 CR8 CR9 CR10 CR11 CR12 CR13 CR14 CR15 CR16 CR17 CR18 CR19 CR20 CR21 CR22 CR23 CR24 CR25 CR26 CR27 CR28 CR29 CR30 CR31 CR32 CR33" CitationID="CR1">1</CitationRef>–<CitationRef CitationID="CR34">34</CitationRef> derived from E. coli. Compared with IMAC and RPC, the purification steps used in this article are more cost-effective and environmentally friendly. This methodology has the potential to be scaled up in an industrial setting.

A novel chromatography method for high purity production of Recombinant human parathyroid hormone from E. coli

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2025年12月30日

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Farhad Mollasoltani,
Milad Adak,
HomaSadat Esfahani,
Elaheh Roshani Yasaghi,
Sirwan Khanchezar

摘要

Human parathyroid hormone (hPTH) shows great potential as a therapeutic option for osteoporosis in patients who do not respond sufficiently to conventional therapies. We utilize recombinant DNA technology to enhance the drug development process and introduce a state-of-the-art purification protocol for the first 34 residues of human parathyroid hormone (hPTH), as these residues are responsible for the majority of its hormonal activity. Typically, the purification of recombinant human parathyroid hormone has relied on His-Tag fusion partners using Ni²⁺-chelating chromatography. This is the first time ion exchange chromatography has been used for the soluble PTH form, and no reports of ion exchange chromatography for this purpose have been documented. SUMO-tag technology and a three-step chromatography process were used to purify recombinant human parathyroid hormone [rhPTH^1–34 on a large scale from Escherichia coli. The rhPTH (1–84) construct was cloned into E. coli host cells and expressed at fermenter scale with a 5-liter volume. The SUMO-hPTH^1–34 fusion protein was extracted and purified using a Capto™ adhere column. The N-terminus of SUMO-hPTH was specifically cleaved with SUMO protease, after which rhPTH was further purified through sequential chromatography on Capto adhere and Capto SP columns. Our purification procedure for rhPTH from E. coli achieved over 98% purity, as confirmed by SDS-PAGE and RP-HPLC. This study demonstrates that the purified rhPTH^1–34 meets the purity standards for commercial drug specifications. We propose that our three-step chromatography process effectively purifies active rhPTH^1–34 derived from E. coli. Compared with IMAC and RPC, the purification steps used in this article are more cost-effective and environmentally friendly. This methodology has the potential to be scaled up in an industrial setting.