NDLI: Expression, purification, and characterisation of nesiritide using an E. coli expression system

Content Provider	Springer Nature Link
Author	Rao, Laxmi S. Niphadkar, Milind P. Paliwal, Dinesh Shekhawat, Rakesh Khare, Aruna G. Uma, S. Thakur, Priti Chutke, Anjali Surlikar, Neelesh Samant, Radhika Zawar, Sagar
Copyright Year	2011
Abstract	Nesiritide, the recombinant human brain natriuretic peptide, is involved in the regulation of cardiovascular homeostasis and has been approved for treatment of patients with congestive heart failure. We prepared a synthetic cDNA construct of Nesiritide to generate a fusion protein with an affinity handle and 41 amino acid peptide of β-galactosidase. The fusion protein was expressed mainly in the inclusion bodies and accounted for approximately 20% of total cellular protein. After purification by Ni-IDA affinity chromatography and renaturation, the fusion protein was cleaved with purified recombinant enterokinase. Nesiritide was purified by pH precipitation/ion exchange chromatography followed by source phenyl chromatography to obtain protein with > 99% purity (determined by RPHPLC) and a mass of 3,464 Daltons. The potency (ED$_{50}$) of the purified protein was equivalent to that of Natrecor (Innovator formulation). Analytical methods were developed to identify oxidised, reduced and other related impurities. The expression strategy described in this work allows the convenient generation of high yield Nesiritide and enabled ease of purification.
Starting Page	688
Ending Page	697
Page Count	10
File Format	PDF
ISSN	12268372
Journal	Biotechnology and Bioprocess Engineering
Volume Number	16
Issue Number	4
e-ISSN	19763816
Language	English
Publisher	The Korean Society for Biotechnology and Bioengineering
Publisher Date	2011-10-05
Publisher Place	Heidelberg
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	brain natriuretic peptide nesiritide expression bioprocess purification characterization Biotechnology Industrial and Production Engineering
Content Type	Text
Resource Type	Article
Subject	Bioengineering Biomedical Engineering Applied Microbiology and Biotechnology Biotechnology

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