Optimized in vitro and in vivo expression of proteorhodopsin: A seven-transmembrane proton pump

Pontus Gourdon, Anna Alfredsson, Anders Pedersen, Erik Malmerberg, Maria Nyblom, Mikael Widell, Ronnie Berntsson, Jarone Pinhassi, Marc Braiman, Örjan Hansson, Nicklas Bonander, Göran Karlsson, Richard Neutze

Research output: Contribution to journalArticle

44 Scopus citations

Abstract

Proteorhodopsin is an integral membrane light-harvesting proton pump that is found in bacteria distributed throughout global surface waters. Here, we present a protocol for functional in vitro production of pR using a commercial cell-free synthesis system yielding 1.0 mg purified protein per milliliter of cell lysate. We also present an optimized protocol for in vivo over-expression of pR in Escherichia coli, and a two-step purification yielding 5 mg of essentially pure functional protein per liter of culture. Both approaches are straightforward, rapid, and easily scalable. Thus either may facilitate the exploitation of pR for commercial biotechnological applications. Finally, the implications of some observations of the in vitro synthesis behavior, as well as preliminary results towards a structural determination of pR are discussed.

Original languageEnglish (US)
Pages (from-to)103-113
Number of pages11
JournalProtein Expression and Purification
Volume58
Issue number1
DOIs
StatePublished - Mar 1 2008

Keywords

  • Cell-free
  • Functional expression
  • Membrane protein
  • Proteorhodopsin

ASJC Scopus subject areas

  • Biotechnology

Fingerprint Dive into the research topics of 'Optimized in vitro and in vivo expression of proteorhodopsin: A seven-transmembrane proton pump'. Together they form a unique fingerprint.

  • Cite this

    Gourdon, P., Alfredsson, A., Pedersen, A., Malmerberg, E., Nyblom, M., Widell, M., Berntsson, R., Pinhassi, J., Braiman, M., Hansson, Ö., Bonander, N., Karlsson, G., & Neutze, R. (2008). Optimized in vitro and in vivo expression of proteorhodopsin: A seven-transmembrane proton pump. Protein Expression and Purification, 58(1), 103-113. https://doi.org/10.1016/j.pep.2007.10.017