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  One-step affinity purification of bacterially produced proteins by means of the "Strep tag" and immobilized recombinant core streptavidin

Schmidt, T. G., & Skerra, A. (1994). One-step affinity purification of bacterially produced proteins by means of the "Strep tag" and immobilized recombinant core streptavidin. Journal of Chromatography A, 676(2), 337-345. doi:10.1016/0021-9673(94)80434-6.

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 Urheber:
Schmidt, Thomas G.M.1, Autor           
Skerra, Arne1, Autor           
Affiliations:
1Department of Molecular Membrane Biology, Max Planck Institute of Biophysics, Max Planck Society, ou_2068290              

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 Zusammenfassung: The “Strep tag” is a nine amino acid peptide with intrinsic streptavidin-binding activity. If this sequence is genetically fused to the C-terminus of a polypeptide the recombinant protein can be directly purified by affinity chromatography from the host cell extract on immobilized streptavidin. However, variations were observed in the suitability of different commercial streptavidin-agarose preparations for this purpose. Therefore, the influence of the source of streptavidin, the coupling chemistry, and the nature of the affinity chromatography resin was investigated. A procedure was developed for the production of recombinant core streptavidin in Escherichia coli, followed by its efficient refolding and purification with an overall yield of up to 140 mg functional protein per 1 1 bacterial culture. When coupled to activated CH-Sepharose 4B this truncated form of streptavidin showed a performance in the affinity chromatography of Strep tag fusion proteins that was superior to all other combinations tested. In contrast to its conventional preparation from Streptomyces strains the recombinant core streptavidin was produced without a proteolytic processing step. Thus, deleterious effects during the streptavidin affinity purification of proteins due to residual proteolytic activity in the immobilized streptavidin were avoided. The versatility of the optimized purification system was demonstrated with five different Strep tag fusion proteins.

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Sprache(n): eng - English
 Datum: 1994-04-191994-02-152001-11-261994-08-05
 Publikationsstatus: Erschienen
 Seiten: 9
 Ort, Verlag, Ausgabe: -
 Inhaltsverzeichnis: -
 Art der Begutachtung: Expertenbegutachtung
 Identifikatoren: DOI: 10.1016/0021-9673(94)80434-6
PMID: 7921186
 Art des Abschluß: -

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Titel: Journal of Chromatography A
Genre der Quelle: Zeitschrift
 Urheber:
Affiliations:
Ort, Verlag, Ausgabe: Amsterdam : Elsevier
Seiten: - Band / Heft: 676 (2) Artikelnummer: - Start- / Endseite: 337 - 345 Identifikator: ISSN: 0021-9673
CoNE: https://pure.mpg.de/cone/journals/resource/954925527837_1