Membrane protein overexpression

    Membrane proteins are notorious for challenges associated with their overexpression. Frequently, overexpression leads to the formation of aggregates or inclusion bodies which is undesirable for most studies. Despite our increasing knowledge on membrane protein biogenesis, at the moment we cannot explain, let alone predict, why few membrane proteins express well but most in fact not. A practical contemporary solution to this problem is broad screening of conditions as expression hosts, fusion proteins, tags and variants or homologs of the membrane protein of interest.

    In addition to this, we make use of tunable promoter systems such as the AraC/PBAD system for E. coli and the nisin promoter in L. lactis. This is relevant as the biogenesis of integral membrane proteins involves many steps such as targeting of the nascent polypeptide to the membrane and insertion into and assembly in the membrane. Each of these steps requires distinct components. Exceeding the capacity of the cell to process the nascent membrane protein correctly will result in the production of aggregated material. By using tunable promoters, we can adjust the expression rate to the capacity of the downstream processing machinery and thereby alleviate some expression difficulties.

    Relevant Publications

    Geertsma ER, Groeneveld M, Slotboom DJ, Poolman B. (2008)
    Quality control of overexpressed membrane proteins.
    Proc Natl Acad Sci U S A. 2008 Apr 15;105(15):5722-7.

    Geertsma ER, Poolman B. (2010)
    Production of membrane proteins in Escherichia coli and Lactococcus lactis.

    Methods Mol Biol. 2010;601:17-38.