Purification of recombinant proteins by chemical removal of the affinity tag.

Détails

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ID Serval
serval:BIB_66D77565FEDE
Type
Article: article d'un périodique ou d'un magazine.
Collection
Publications
Institution
Titre
Purification of recombinant proteins by chemical removal of the affinity tag.
Périodique
Applied Biochemistry and Biotechnology
Auteur⸱e⸱s
Rais-Beghdadi C., Roggero M.A., Fasel N., Reymond C.D.
ISSN
0273-2289 (Print)
ISSN-L
0273-2289
Statut éditorial
Publié
Date de publication
1998
Volume
74
Numéro
2
Pages
95-103
Langue
anglais
Résumé
The efficient removal of a N- or C-terminal purification tag from a fusion protein is necessary to obtain a protein in a pure and active form, ready for use in human or animal medicine. Current techniques based on enzymatic cleavage are expensive and result in the presence of additional amino acids at either end of the proteins, as well as contaminating proteases in the preparation. Here we evaluate an alternative method to the one-step affinity/protease purification process for large-scale purification. It is based upon the cyanogen bromide (CNBr) cleavage at a single methionine placed in between a histidine tag and a Plasmodium falciparum antigen. The C-terminal segment of the circumsporozoite polypeptide was expressed as a fusion protein with a histidine tag in Escherichia coli purified by Ni-NAT agarose column chromatography and subsequently cleaved by CNBr to obtain a polypeptide without any extraneous amino acids derived from the cleavage site or from the affinity purification tag. Thus, a recombinant protein is produced without the need for further purification, demonstrating that CNBr cleavage is a precise, efficient, and low-cost alternative to enzymatic digestion, and can be applied to large-scale preparations of recombinant proteins.
Mots-clé
Amino Acid Sequence, Animals, Chromatography, Affinity, Cyanogen Bromide/chemistry, Escherichia coli, Histidine/chemistry, Mass Spectrometry, Molecular Sequence Data, Plasmodium falciparum, Protozoan Proteins/biosynthesis, Protozoan Proteins/chemistry, Recombinant Proteins/biosynthesis, Recombinant Proteins/isolation &amp, purification
Pubmed
Web of science
Open Access
Oui
Création de la notice
24/01/2008 15:02
Dernière modification de la notice
14/02/2022 7:55
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