Format

Send to

Choose Destination
See comment in PubMed Commons below
J Struct Biol. 2011 Aug;175(2):178-88. doi: 10.1016/j.jsb.2011.02.001. Epub 2011 Feb 12.

Deciphering correct strategies for multiprotein complex assembly by co-expression: application to complexes as large as the histone octamer.

Author information

1
Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Département de Biologie Intégrative, Institut National de Santé et de Recherche Médicale (INSERM) U964/Centre National de Recherche Scientifique (CNRS) UMR 1704/Université de Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch, France.

Abstract

Macromolecular complexes are responsible for most of the essential mechanisms in cells, leading to a broad interest in their purification and characterization. Co-expression is now widely recognized as a major technique for assembling multiprotein complexes and many co-expression systems are currently available for performing co-expression experiments in different hosts. However, comparative knowledge on co-expression strategies is still crucially lacking. Using versatile co-expression systems for Escherichia coli, the pET-MCN and pET-MCP series, and ternary protein complexes as examples, we demonstrate how to successfully delineate correct co-expression strategies. Specifically, an appropriate, complex-dependent approach alleviates stoichiometry imbalance and yield problems, and even failure in producing complexes. Importantly, some of the parameters influencing co-expression strategies appear independent of the expression host, thus having implications for co-expression in eukaryotic hosts. By further using these strategies, we show that co-expression in E. coli enables reconstitution of protein complexes as large as the deubiquitination module of the SAGA transcription factor and the histone octamer.

PMID:
21320604
DOI:
10.1016/j.jsb.2011.02.001
[Indexed for MEDLINE]
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for Elsevier Science
    Loading ...
    Support Center