Use of Ssp dnaB mini-intein as a fusion partner for preparation of recombinant human brain natriuretic peptide.
- Author:
Lie DI
1
;
Hong-Wei ZHANG
;
Lang-Lai XU
Author Information
1. College of Life Sciences, Nanjing Agricultural University, Nanjing 210095, China.
- Publication Type:Journal Article
- MeSH:
Animals;
DnaB Helicases;
biosynthesis;
genetics;
Escherichia coli;
genetics;
metabolism;
Genetic Vectors;
Inteins;
genetics;
Natriuretic Peptide, Brain;
biosynthesis;
genetics;
Protein Engineering;
methods;
Protein Splicing;
Rabbits;
Recombinant Fusion Proteins;
biosynthesis;
genetics;
pharmacology
- From:
Chinese Journal of Biotechnology
2006;22(2):180-203 passim
- CountryChina
- Language:Chinese
-
Abstract:
Human brain natriuretic peptide (hBNP) was used clinically for the treatment of acute decompensated congestive heart failure. In this paper, hBNP was expressed as a fusion protein with a histidine tag and Ssp dnaB mini-intein which was capable of self-cleavage. After affinity chromatography with Ni-Sepharose and renaturation, the fusion protein was enriched with CM-cellulose. Ssp dnaB mini-intein mediated peptide-bond hydrolysis was triggered by shifting the pH and temperature in the CM-cellulose column, which let to the release of hBNP from the fusion protein and the separation of hBNP from His-DnaB. The hBNP sample was further purified by C4 reverse phase HPLC, and 2.8mg of the peptide with homogeneity of 97% was obtained from one liter of culture medium. The biological activity was assayed in vitro, which indicated that hBNP had a potent vasodilatory effect on rabbit aortic strips with an EC50 of 1.94 x 10(-6) mg/mL.
