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  Protein complex purification from Thermoplasma acidophilum using a phage display library

Hubert, A., Mitani, Y., Tamura, T., Boicu, M., & Nagy, I. (2014). Protein complex purification from Thermoplasma acidophilum using a phage display library. JOURNAL OF MICROBIOLOGICAL METHODS, 98, 15-22. doi:10.1016/j.mimet.2013.12.010.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0019-8B67-C Version Permalink: http://hdl.handle.net/11858/00-001M-0000-0019-8B68-A
Genre: Journal Article

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 Creators:
Hubert, Agnes1, Author              
Mitani, Yasuo2, Author
Tamura, Tomohiro2, Author
Boicu, Marius1, Author              
Nagy, Istvan1, Author              
Affiliations:
1Baumeister, Wolfgang / Molecular Structural Biology, Max Planck Institute of Biochemistry, Max Planck Society, ou_1565142              
2external, ou_persistent22              

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Free keywords: CDC48/P97 ATPASE HOMOLOG; TRICORN PROTEASE; VAT; PROTEOMICS; GELSProtein complex; Phage display library; Thermoplasma acidophilum; Electron microscopy;
 Abstract: We developed a novel protein complex isolation method using a single-chain variable fragment (scFv) based phage display library in a two-step purification procedure. We adapted the antibody-based phage display technology which has been developed for single target proteins to a protein mixture containing about 300 proteins, mostly subunits of Thermoplasma acidophilum complexes. T. acidophilum protein specific phages were selected and corresponding scFvs were expressed in Escherichia coli. E. coil cell lysate containing the expressed His-tagged scFv specific against one antigen protein and T. acidophilum crude cell lysate containing intact target protein complexes were mixed, incubated and subjected to protein purification using affinity and size exclusion chromatography steps. This method was confirmed to isolate intact particles of thermosome and proteasome suitable for electron microscopy analysis and provides a novel protein complex isolation strategy applicable to organisms where no genetic tools are available. (C) 2013 Elsevier B.V. All rights reserved.

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Language(s): eng - English
 Dates: 2014-03
 Publication Status: Published in print
 Pages: 8
 Publishing info: -
 Table of Contents: -
 Rev. Method: Peer
 Degree: -

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Title: JOURNAL OF MICROBIOLOGICAL METHODS
Source Genre: Journal
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Publ. Info: PO BOX 211, 1000 AE AMSTERDAM, NETHERLANDS : ELSEVIER SCIENCE BV
Pages: - Volume / Issue: 98 Sequence Number: - Start / End Page: 15 - 22 Identifier: ISSN: 0167-7012