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Title 

Novel vectors for the convenient cloning and expression of In vivo biotinylated proteins in Escherichia coli

Authors 

Eun Wie ChoJung Hyun ParkShin Young NaKil Lyong Kim

Publisher 

Springer Verlag (Germany)

Issue Date 

1999

Citation 

Journal of Biochemistry and Molecular Biology, vol. 32, no. 5, pp. 497-501

Keywords 

In vivo biotinylationFusion proteinTGF-βThioredoxin

Abstract 

Biotinylation of recombinant proteins is a powerful tool for the detection and analysis of proteins of interest in a large variety of assay systems. The recent development of in vivo biotinylation techniques in E. coli has opened new possibilities for the production of site-specifically biotinylated proteins without the need for further manipulation after the isolation of the recombinantly expressed proteins. In the present study, a novel vector set was generated which allows the convenient cloning and expression of proteins of interest fused with an N-terminal in vivo biotinylated thioredoxin (TRX) protein. These vectors were derived from the previously reported pBIOTRX vector into which was incorporated part of the pBluescript II+ phagemid multiple cloning site (MCS), amplified by PCR using a pair of sophisticated oligonucleotide primers. The functionality of these novel vectors was examined in this system by recombinant expression of rat transforming growth factor-β. Western-blot analysis using TRX-specific antibodies or peroxidase- conjugated streptavidin confirmed the successful induction of the fusion protein and the in vivo conjugation of biotin molecules, respectively. The convenience of molecular subcloning provided by the MCS and the effective in vivo biotinylation of proteins of interest makes this novel vector set an interesting alternative for the production of biotinylated proteins.

ISSN 

1225-8687

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2017-04-19


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