Characterization and Expression Profiling of Recombinant Parathyroid Hormone (rhPTH) Analog 1–34 in Escherichia coli, Precise with Enhanced Biological Activity
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Human truncated parathyroid hormone [hPTH (1–34)], a peptide hormone which accelerated the research interest towards the theraputical applications. This study outlines the effective expression of [hPTH (1–34)] in Escherichia coli and the recuperation of highly soluble truncated PTH from the fusion protein by proteolytic digestion. Successful expression of glutathione S-transferase (GST) fusion protein was achieved by incorporating truncated PTH with an enzymatic cleavage site into the “N” terminal GST of pGEX-4T-3 expression vector. Under the optimized condition, we achieved more than 120 mg of pure hPTH (1–34) per liter of bacterial culture, with an overall yield of 39%. Purification process was carried out through immobilized metal ion chromatography and membrane filter to produce maximum purity. Physical characterization using western blot analysis showed that the extracted truncated PTH is intact and reacts with anti-PTH antibodies. In vitro analysis of PTH stimulated adenylyl cyclase activation in UMR 106 cells confirmed biological activity for purified protein. We believe this vector and production methodology represents a generally applicable tool for the generation of recombinant peptides.
KeywordsTruncated parathyroid hormone Escherichia coli Glutathione S-transferase Enterokinase Purification
Authors thank Acharya Nagarjuna University, Andhra Pradesh for providing all facilities required for this study.
This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors.
Compliance with Ethical Standards
Conflict of interest
Rajeenkanna Chilakapati and Chanchal Thomas Mannully carried out this experiment. Mrinmoy Ghosh arrange the data and stastical presentation. K.K. Pulicherla planned the experiment. There was no conflict of interest among the authors.
Research Involving Human and Animal Participants
This article does not contain any studies with human participants or animals performed by any of the authors.
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