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Hyderabad, India

Gangireddy S.R.,Virchow Research Institute | Gangireddy S.R.,Acharya Nagarjuna University | Madhavi R.D.,Virchow Research Institute | Ravikanth K.R.,Virchow Research Institute | And 4 more authors.
Current Trends in Biotechnology and Pharmacy | Year: 2010

Human parathyroid hormone (hPTH), synthesized in the parathyroid gland as a linear peptide, is one of the key regulatory molecules in calcium homeostasis and bone resorption. The N terminus region of hPTH (1-34) is a functionally important part of the molecule that is sufficient and necessary for potently executing most of the hormonal actions. Therefore, hPTH (1-34) is considered to be an attractive therapeutic agent in the treatment of osteoporosis. Here, we describe a high yield expression and purification method for the production of hPTH (1-34) from Escherichia coli. The hPTH (1-34) was expressed as a fusion protein in soluble form, and found to be more than 25% of total proteins. The fusion protein was first purified by GST affinity chromatography and, after cleavage of GST with Factor Xa, the peptide (hPTH 1-34) was further purified by reversed phase Source-30™ chromatography. This double purification strategy produced 30mg/l of hPTH (1-34) with purity = 98%. The identity of the purified peptide was confirmed by mass spectrometry and N-terminal sequencing analysis. The biological activity of the peptide was confirmed in the rat osteogenic cell line UMR-106 by measuring cAMP levels, which were identical to hPTH standards, indicating that purified rhPTH (1-34) has full biological activity.

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