His-tagged yeast 6-phosphofructo-2-kinase was overexpressed in the yeast strain DFY658 under the control of the Gal1 promoter. Here we describe a simple and fast purification protocol for the recombinant enzyme under native conditions using a HiTrap affinity column loaded with CuSO(4). The use of MALDI-TOF MS after in-gel-digestion enabled us to identify a critical contamination of the end product as yeast alcohol dehydrogenase1 (Adh1p). After identification this contaminant could be efficiently removed by carrying out the washing steps at 25 degrees C instead of at 4 degrees C. To reduce the cellular proteolytic activities a low phosphate concentration in the growth medium was applied. This simple modification of the yeast cell growth conditions increased significantly the yield of the recombinant protein.
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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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