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Expression of proteins using the third domain of the Escherichia coli periplasmic-protein TolA as a fusion partner

Lookup NU author(s): Isa Gokce, Professor Jeremy Lakey

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Abstract

The third domain of the periplasmic protein TolA from Escherichia coli (TolAIII) was used as a fusion partner in the expression of various proteins from bacteria and eukaryotes. TolAIII is small domain, expressed in high yields as a soluble protein in the cytoplasm of E. coli. Proteins were linked to the C-terminus of TolAIII by a short flexible linker containing sites for endopeptidases. Three different vectors were prepared, containing sites for enterokinase, thrombin or factor Xa. Fusion proteins also contain a His6-Ser2 tag at their N-terminus for easier purification. Up to 90 mg fusion protein per liter bacterial culture was obtained using these vectors. Colicin N R-domain was expressed with this system as a fusion and processed further for functional studies. The yield of final pure R-domain was doubled as compared to the direct expression. The system may prove to be useful in the preparation of other peptides and proteins. © 2002 Elsevier Science (USA). All rights reserved.


Publication metadata

Author(s): Anderluh G, Gokce I, Lakey JH

Publication type: Article

Publication status: Published

Journal: Protein Expression and Purification

Year: 2003

Volume: 28

Issue: 1

Pages: 173-181

ISSN (print): 1046-5928

ISSN (electronic): 1096-0279

Publisher: Academic Press

URL: http://dx.doi.org/10.1016/S1046-5928(02)00681-2

DOI: 10.1016/S1046-5928(02)00681-2

PubMed id: 12651122


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