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    Simultaneous recording from four lipid bilayers

2021 - Saccharomyces cerevisiae as a superior host for overproduction of prokaryotic integral membrane proteins

Icon Orbit Mini  Orbit Mini Publication in Current Research in Structural Biology (2021)

Spruce Preisler S., Wiuf A.D., Friss M., Kjaergaard L., Hurd M., Ramos Becares E., Nurup C.N., Bjoerkskov F.B., Szathmary Z., Gourson P.E., Calloe K., Klaerke D.A., Gotfryd K., Pedersen P.A.

Current Research in Structural Biology (2021) doi: 10.1016/j.crstbi.2021.02.001.


Integral membrane proteins (IMPs) constitute ~30% of all proteins encoded by the genome of any organism and Escherichia coli remains the first-choice host for recombinant production of prokaryotic IMPs. However, the expression levels of prokaryotic IMPs delivered by this bacterium are often low and overproduced targets often accumulate in inclusion bodies. The targets are therefore often discarded to avoid an additional and inconvenient refolding step in the purification protocol. Here we compared expression of five prokaryotic (bacterial and archaeal) IMP families in E. coli and Saccharomyces cerevisiae. We demonstrate that our S. cerevisiae-based production platform is superior in expression of four investigated IMPs, overall being able to deliver high quantities of active target proteins. Surprisingly, in case of the family of zinc transporters (Zrt/Irt-like proteins, ZIPs), S. cerevisiae rescued protein expression that was undetectable in E. coli. We also demonstrate the effect of localization of the fusion tag on expression yield and sample quality in detergent micelles. Lastly, we present a road map to achieve the most efficient expression of prokaryotic IMPs in our yeast platform. Our findings demonstrate the great potential of S. cerevisiae as host for high-throughput recombinant overproduction of bacterial and archaeal IMPs for downstream biophysical characterization.

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