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Related Experiment Video

Updated: Jun 26, 2026

Green Fluorescent Protein-based Expression Screening of Membrane Proteins in Escherichia coli
08:46

Green Fluorescent Protein-based Expression Screening of Membrane Proteins in Escherichia coli

Published on: January 6, 2015

Genetic selection system for improving recombinant membrane protein expression in E. coli.

Elizabeth Massey-Gendel1, Anni Zhao, Gabriella Boulting

  • 1Department of Chemistry and Biochemistry, University of California, Los Angeles, USA.

Protein Science : a Publication of the Protein Society
|January 24, 2009
PubMed
Summary

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Researchers developed a novel selection strategy to isolate Escherichia coli mutants, termed EXP strains, that significantly enhance membrane protein expression. This method links protein production to survival, enabling up to 75-fold increases in recombinant membrane protein yields.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Structural Biology

Background:

  • Low yield in recombinant expression poses a significant challenge for membrane protein characterization and structure determination.
  • Developing efficient methods for increasing membrane protein expression is crucial for structural and functional studies.

Purpose of the Study:

  • To design and implement a selection strategy for isolating mutant Escherichia coli strains with improved membrane protein expression.
  • To establish a rapid method for plasmid curing in selected strains.

Main Methods:

  • A selection strategy was developed by fusing the membrane protein of interest to a C-terminal selectable marker.
  • Survival on selective media was directly linked to the expression of the targeted membrane protein.

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Last Updated: Jun 26, 2026

Green Fluorescent Protein-based Expression Screening of Membrane Proteins in Escherichia coli
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Green Fluorescent Protein-based Expression Screening of Membrane Proteins in Escherichia coli

Published on: January 6, 2015

Characterization of Membrane Transporters by Heterologous Expression in E. coli and Production of Membrane Vesicles
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Characterization of Membrane Transporters by Heterologous Expression in E. coli and Production of Membrane Vesicles

Published on: December 31, 2019

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  • A plasmid curing method using the homing endonuclease I-CreI was introduced.

    • Main Results:

    • The selection system successfully isolated mutant strains (EXP strains) exhibiting up to 75-fold increased expression of a target membrane protein (Rv1337).
    • The developed EXP strains also demonstrated improved expression for other unrelated membrane proteins, with one showing a roughly 90-fold increase.
    • The rapid plasmid curing method efficiently removed selection plasmids from isolated strains.

    Conclusions:

    • The developed selection strategy is effective in isolating bacterial strains with significantly enhanced membrane protein expression.
    • This approach provides a valuable tool for overcoming expression barriers in membrane protein research.
    • The EXP strains offer a promising resource for future structural and functional investigations of membrane proteins.