Efficient production of membrane-integrated and detergent-soluble G protein-coupled receptors in Escherichia coli

Authors

  • A. James Link,

    1. Department of Chemical Engineering, The University of Texas at Austin, Austin, Texas 78712, USA
    2. Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin, Texas 78712, USA
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    • Present address: Department of Chemical Engineering, Princeton University, Princeton, NJ 08544, USA.

  • Georgios Skretas,

    1. Department of Chemical Engineering, The University of Texas at Austin, Austin, Texas 78712, USA
    2. Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin, Texas 78712, USA
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  • Eva-Maria Strauch,

    1. Department of Chemistry and Biochemistry, The University of Texas at Austin, Austin, Texas 78712, USA
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  • Nandini S. Chari,

    1. Department of Chemistry and Biochemistry, The University of Texas at Austin, Austin, Texas 78712, USA
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  • George Georgiou

    Corresponding author
    1. Department of Chemical Engineering, The University of Texas at Austin, Austin, Texas 78712, USA
    2. Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin, Texas 78712, USA
    3. Department of Biomedical Engineering, The University of Texas at Austin, Austin, Texas 78712, USA
    4. Department of Molecular Genetics and Microbiology, The University of Texas at Austin, Austin, Texas 78712, USA
    • Department of Chemical Engineering, University of Texas at Austin, 1 University Station C0400, Austin, TX 78712-0231, USA; fax: (512) 471-7963.
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Abstract

G protein-coupled receptors (GPCRs) are notoriously difficult to express, particularly in microbial systems. Using GPCR fusions with the green fluorescent protein (GFP), we conducted studies to identify bacterial host effector genes that result in a general and significant enhancement in the amount of membrane-integrated human GPCRs that can be produced in Escherichia coli. We show that coexpression of the membrane-bound AAA+ protease FtsH greatly enhances the expression yield of four different class I GPCRs, irrespective of the presence of GFP. Using this new expression system, we produced 0.5 and 2 mg/L of detergent-solubilized and purified full-length central cannabinoid receptor (CB1) and bradykinin receptor 2 (BR2) in shake flask cultures, respectively, two proteins that had previously eluded expression in microbial systems.

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