ArticleProtein science : a publication of the Protein Society2014
Method to increase the yield of eukaryotic membrane protein expression in Saccharomyces cerevisiae for structural and functional studies.
Article in Protein science : a publication of the Protein Society, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 18 papers.
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18 citing papers in PubMed.
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- Programmable Lipid Bilayer Tension-Control Apparatus for Quantitative Mechanobiology.ACS nano · 2024Article
- Identification of SLC45A4 as a pain gene encoding a neuronal polyamine transporter.bioRxiv : the preprint server for biology · 2024Article
- Lysosomal membrane transporter purification and reconstitution for functional studies.Molecular biology of the cell · 2024Article
- A Mitosome With Distinct Metabolism in the Uncultured Protist Parasite Paramikrocytos canceri (Rhizaria, Ascetosporea).Genome biology and evolution · 2023Article
- Structural basis for proton coupled cystine transport by cystinosin.Nature communications · 2022Article
- Heterologous Expression of Full-Length and Truncated Human ZIP4 Zinc Transporter inBiomolecules · 2022Article
- The high-throughput production of membrane proteins.Emerging topics in life sciences · 2021Article
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- An improved fluorescent tag and its nanobodies for membrane protein expression, stability assay, and purification.Communications biology · 2020Article
- Changes in Membrane Protein Structural Biology.Biology · 2020Review
- An Improved Strategy for Fluorescent Tagging of Membrane Proteins for Overexpression and Purification in Mammalian Cells.Biochemistry · 2018Article
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- Expression and Subcellular Distribution of GFP-Tagged Human Tetraspanin Proteins in Saccharomyces cerevisiae.PloS one · 2015Article
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2 authors.
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Abstract
Despite recent successes in the structure determination of eukaryotic membrane proteins, the total number of structures of these important proteins is severely underrepresented in the Protein Data Bank. Although prokaryotic homologues provide valuable mechanistic insight, they often lack crucial details, such as post-translational modification and additional intra or extracellular domains that are important for understanding the function and regulation of these proteins in eukaryotic cells. The production of milligram quantities of recombinant protein is still a serious obstacle to the structural and functional characterization of these proteins. Here, we report a modification to a previously described over expression system using the simple eukaryote Saccharomyces cerevisiae that can increase overall protein yield and improve downstream purification procedures. Using a metabolic marker under the control of a truncated promoter, we show that expression levels for several membrane transporters are increased fourfold. We further demonstrate that the increase in expression for our test proteins resulted in a concomitant increase in functional protein. Using this system, we were able to increase the expression level of a plant transporter, NRT1.1, which was a key factor in its structural and functional characterization.
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