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Open Access Green as soon as Postprint is submitted to ZB.
High-throughput protein expression screening and purification in Escherichia coli.
Methods 55, 65-72 (2011)
Escherichia coli (E. coli) is the most widely used expression system for the production of recombinant proteins for structural and functional studies. However, to obtain milligrams of soluble proteins is still challenging since many proteins are expressed in an insoluble form without optimization. Therefore when working with tens of proteins or protein domains it is recommended that high-throughput expression screening at a small scale (1-4ml of culture) is carried out to identify the optimal conditions for soluble protein production. Once determined, these culture conditions can be applied at a large scale to produce sufficient protein for structural or functional studies. We describe a procedure that has enabled the systematic screening of culture conditions or fusion-tags on hundreds of cultures per week. The analysis of the optimal conditions for the soluble production of these proteins helped us to design a simple and efficient protocol for soluble protein expression screening. This protocol has since been used on hundreds of proteins and is illustrated with the genome wide scale production of proteins containing the DNA binding domains of Ciona intestinalis.
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Publication type
Article: Journal article
Document type
Scientific Article
Keywords
High-throughput; Solubility screening; Escherichia coli; Thioredoxin; Ciona intestinalis
ISSN (print) / ISBN
1046-2023
e-ISSN
1095-9130
Journal
Methods
Quellenangaben
Volume: 55,
Issue: 1,
Pages: 65-72
Publisher
Elsevier
Publishing Place
Amsterdam [u.a.]
Non-patent literature
Publications
Reviewing status
Peer reviewed
Institute(s)
Institute of Structural Biology (STB)