Expression of protein complexes using multiple Escherichia coli protein co-expression systems: a benchmarking study.

2.50
Hdl Handle:
http://hdl.handle.net/10033/208749
Title:
Expression of protein complexes using multiple Escherichia coli protein co-expression systems: a benchmarking study.
Authors:
Busso, Didier; Peleg, Yoav; Heidebrecht, Tatjana; Romier, Christophe; Jacobovitch, Yossi; Dantes, Ada; Salim, Loubna; Troesch, Edouard; Schuetz, Anja; Heinemann, Udo; Folkers, Gert E; Geerlof, Arie; Wilmanns, Matthias; Polewacz, Andrea; Quedenau, Claudia; Büssow, Konrad ( 0000-0003-2031-5942 ) ; Adamson, Rachel; Blagova, Elena; Walton, Julia; Cartwright, Jared L; Bird, Louise E; Owens, Raymond J; Berrow, Nick S; Wilson, Keith S; Sussman, Joel L; Perrakis, Anastassis; Celie, Patrick H N
Abstract:
Escherichia coli (E. coli) remains the most commonly used host for recombinant protein expression. It is well known that a variety of experimental factors influence the protein production level as well as the solubility profile of over-expressed proteins. This becomes increasingly important for optimizing production of protein complexes using co-expression strategies. In this study, we focus on the effect of the choice of the expression vector system: by standardizing experimental factors including bacterial strain, cultivation temperature and growth medium composition, we compare the effectiveness of expression technologies used by the partners of the Structural Proteomics in Europe 2 (SPINE2-complexes) consortium. Four different protein complexes, including three binary and one ternary complex, all known to be produced in the soluble form in E. coli, are used as the benchmark targets. The respective genes were cloned by each partner into their preferred set of vectors. The resulting constructs were then used for comparative co-expression analysis done in parallel and under identical conditions at a single site. Our data show that multiple strategies can be applied for the expression of protein complexes in high yield. While there is no 'silver bullet' approach that was infallible even for this small test set, our observations are useful as a guideline to delineate co-expression strategies for particular protein complexes.
Affiliation:
Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Institut National de Santé et de Recherche Médicale (Inserm), U964/Centre National deRecherche Scientifique (CNRS), UMR 7104, Université de Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch, France.
Citation:
Expression of protein complexes using multiple Escherichia coli protein co-expression systems: a benchmarking study. 2011, 175 (2):159-70 J. Struct. Biol.
Journal:
Journal of structural biology
Issue Date:
Aug-2011
URI:
http://hdl.handle.net/10033/208749
DOI:
10.1016/j.jsb.2011.03.004
PubMed ID:
21382497
Type:
Article
Language:
en
ISSN:
1095-8657
Appears in Collections:
Publications from Division of Molekulare Struktur Biologie (MOSB)

Full metadata record

DC FieldValue Language
dc.contributor.authorBusso, Didieren
dc.contributor.authorPeleg, Yoaven
dc.contributor.authorHeidebrecht, Tatjanaen
dc.contributor.authorRomier, Christopheen
dc.contributor.authorJacobovitch, Yossien
dc.contributor.authorDantes, Adaen
dc.contributor.authorSalim, Loubnaen
dc.contributor.authorTroesch, Edouarden
dc.contributor.authorSchuetz, Anjaen
dc.contributor.authorHeinemann, Udoen
dc.contributor.authorFolkers, Gert Een
dc.contributor.authorGeerlof, Arieen
dc.contributor.authorWilmanns, Matthiasen
dc.contributor.authorPolewacz, Andreaen
dc.contributor.authorQuedenau, Claudiaen
dc.contributor.authorBüssow, Konraden
dc.contributor.authorAdamson, Rachelen
dc.contributor.authorBlagova, Elenaen
dc.contributor.authorWalton, Juliaen
dc.contributor.authorCartwright, Jared Len
dc.contributor.authorBird, Louise Een
dc.contributor.authorOwens, Raymond Jen
dc.contributor.authorBerrow, Nick Sen
dc.contributor.authorWilson, Keith Sen
dc.contributor.authorSussman, Joel Len
dc.contributor.authorPerrakis, Anastassisen
dc.contributor.authorCelie, Patrick H Nen
dc.date.accessioned2012-02-03T12:10:12Zen
dc.date.available2012-02-03T12:10:12Zen
dc.date.issued2011-08en
dc.identifier.citationExpression of protein complexes using multiple Escherichia coli protein co-expression systems: a benchmarking study. 2011, 175 (2):159-70 J. Struct. Biol.en
dc.identifier.issn1095-8657en
dc.identifier.pmid21382497en
dc.identifier.doi10.1016/j.jsb.2011.03.004en
dc.identifier.urihttp://hdl.handle.net/10033/208749en
dc.description.abstractEscherichia coli (E. coli) remains the most commonly used host for recombinant protein expression. It is well known that a variety of experimental factors influence the protein production level as well as the solubility profile of over-expressed proteins. This becomes increasingly important for optimizing production of protein complexes using co-expression strategies. In this study, we focus on the effect of the choice of the expression vector system: by standardizing experimental factors including bacterial strain, cultivation temperature and growth medium composition, we compare the effectiveness of expression technologies used by the partners of the Structural Proteomics in Europe 2 (SPINE2-complexes) consortium. Four different protein complexes, including three binary and one ternary complex, all known to be produced in the soluble form in E. coli, are used as the benchmark targets. The respective genes were cloned by each partner into their preferred set of vectors. The resulting constructs were then used for comparative co-expression analysis done in parallel and under identical conditions at a single site. Our data show that multiple strategies can be applied for the expression of protein complexes in high yield. While there is no 'silver bullet' approach that was infallible even for this small test set, our observations are useful as a guideline to delineate co-expression strategies for particular protein complexes.en
dc.language.isoenen
dc.subject.meshAcademies and Institutesen
dc.subject.meshCCAAT-Binding Factoren
dc.subject.meshCell Cycle Proteinsen
dc.subject.meshCloning, Molecularen
dc.subject.meshEscherichia colien
dc.subject.meshEuropeen
dc.subject.meshGenetic Vectorsen
dc.subject.meshInternational Cooperationen
dc.subject.meshIsraelen
dc.subject.meshMultiprotein Complexesen
dc.subject.meshRecombinant Proteinsen
dc.subject.meshTranscription Factors, TFIIen
dc.titleExpression of protein complexes using multiple Escherichia coli protein co-expression systems: a benchmarking study.en
dc.typeArticleen
dc.contributor.departmentInstitut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Institut National de Santé et de Recherche Médicale (Inserm), U964/Centre National deRecherche Scientifique (CNRS), UMR 7104, Université de Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch, France.en
dc.identifier.journalJournal of structural biologyen

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