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

Item Type:	Article
Title:	Expression of protein complexes using multiple E. coli protein co-expression systems: a benchmarking study
Creators Name:	Busso, D. and Peleg, Y. and Heidebrecht, T. and Romier, C. and Jacobovitch, Y. and Dantes, A. and Salim, L. and Troesch, E. and Schuetz, A. and Heinemann, U. and Folkers, G.E. and Geerlof, A. and Wilmanns, M. and Polewacz, A. and Quedenau, C. and Büssow, K. and Adamson, R. and Blagova, E. and Walton, J. and Cartwright, J.L. and Bird, L.E. and Owens, R.J. and Berrow, N.S. and Wilson, K.S. and Sussman, J.L. and Perrakis, A. and Celie, P.H.
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.
Keywords:	Escherichia Coli, Co-Expression, Cloning Strategies, Enzyme-Free Cloning, Gateway(TM), In-Fusion(TM), LIC, Restriction-Free Cloning
Source:	Journal of Structural Biology
ISSN:	1047-8477
Publisher:	Academic Press
Volume:	175
Number:	2
Page Range:	159-170
Date:	August 2011
Official Publication:	https://doi.org/10.1016/j.jsb.2011.03.004
PubMed:	View item in PubMed

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