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Eukaryotic integral membrane protein expression utilizing the Escherichia coli glycerol-conducting channel protein (GlpF)

Abstract

A fusion protein expression system is described that allows for production of eukaryotic integral membrane proteins in Escherichia coli (E. coli). The eukaryotic membrane protein targets are fused to the C terminus of the highly expressed E. coli inner membrane protein, GlpF (the glycerol-conducting channel protein). The generic utility of this system for heterologous membrane-protein expression is demonstrated by the expression and insertion into the E. coli cell membrane of the human membrane proteins: occludin, claudin 4, duodenal ferric reductase and a J-type inwardly rectifying potassium channel. The proteins are produced with C-terminal hexahistidine tags (to permit purification of the expressed fusion proteins using immobilized metal affinity chromatography) and a peptidase cleavage site (to allow recovery of the unfused eukaryotic protein).

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Acknowledgements

This work was supported by an award from the Heptagon Fund. Special thanks go to Prof. Robert Hider and Drs Tom Seddon, Sophie Khanna, Ian Hill, Jon Oakhill and Colin Dolphin.

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Correspondence to David Barlow.

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Neophytou, I., Harvey, R., Lawrence, J. et al. Eukaryotic integral membrane protein expression utilizing the Escherichia coli glycerol-conducting channel protein (GlpF). Appl Microbiol Biotechnol 77, 375–381 (2007). https://doi.org/10.1007/s00253-007-1174-7

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Keywords

  • Recombinant protein
  • Membrane protein
  • Gene expression
  • Protein engineering
  • Escherichia coli