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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Evidence ID | Analyze ID | Gene/Complex | Systematic Name/Complex Accession | Qualifier | Gene Ontology Term ID | Gene Ontology Term | Aspect | Annotation Extension | Evidence | Method | Source | Assigned On | Reference |
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Evidence ID | Analyze ID | Gene | Gene Systematic Name | Phenotype | Experiment Type | Experiment Type Category | Mutant Information | Strain Background | Chemical | Details | Reference |
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Evidence ID | Analyze ID | Gene | Gene Systematic Name | Disease Ontology Term | Disease Ontology Term ID | Qualifier | Evidence | Method | Source | Assigned On | Reference |
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Evidence ID | Analyze ID | Regulator | Regulator Systematic Name | Target | Target Systematic Name | Direction | Regulation of | Happens During | Regulator Type | Direction | Regulation Of | Happens During | Method | Evidence | Strain Background | Reference |
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Site | Modification | Modifier | Source | Reference |
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Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Allele | Assay | Annotation | Action | Phenotype | SGA score | P-value | Source | Reference | Note |
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Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Assay | Annotation | Action | Modification | Source | Reference | Note |
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Complement ID | Locus ID | Gene | Species | Gene ID | Strain background | Direction | Details | Source | Reference |
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Evidence ID | Analyze ID | Dataset | Description | Keywords | Number of Conditions | Reference |
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Evidence ID | Analyze ID | File | Description |
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