Economically feasible bioconversion of lignocelluloses into fuels and chemicals is dependent on efficient utilization of all available sugars in lignocellulosic biomass, including hextose and pentose. Previously, we constructed a xylose fermenting strain of Saccharomyces cerevisiae through metabolic engineering and enhanced its xylose utilization capability through evolutionary engineering. However, the key mechanism of improved xylose utilization and xylose isomerase activity was not identified. In this study, we applied the concept of inverse metabolic engineering to identify the factors involved in improving xylose utilization. Genomic sequencing of the evolved strain with fast xylose utilization capability was performed, and mutations possibly affecting xylose utilization were screened. Further genetic analysis of these mutant genes revealed that mutations in ASK10 (both the site-directed mutation ASK10M475R as well as ASK10 deletion), a stress response regulator-encoding gene, improved growth on xylose and enhanced xylose isomerase activity. We found that mutation of Ask10p did not increase xylose isomerase activity through interacting with the xylose isomerase protein or through directly regulating the xylA gene transcription. Although ASK10 deletion increased the copy number of the plasmid and improved the transcription of xylA, the site-direct mutation ASK10M475R did not change the plasmid copy number. Interestingly, we found that both the site-directed mutation ASK10M475R and ASK10 deletion up-regulated the transcription of molecular chaperone-encoding genes HSP26, SSA1 and HSP104, thereby facilitating the protein folding of xylose isomerase and enhancing xylose isomerase activity. This study revealed the important mechanism of chaperones in xylose isomerase activity regulation, and it provides valuable insights for efficient xylose metabolic strain development.
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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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