Lacto-N-neotetraose (LNnT) is one of the important ingredients of human milk oligosaccharides, with various promising health effects for infants. In this study, the biosynthetic pathway of LNnT was constituted in GRAS (generally recognized as safe) host Saccharomyces cerevisiae by coexpressing β1,3-N-acetylglucosaminyltransferase (LgtA), β1,4-galactosyltransferase (LgtB), lactose permease (Lac12), and UDP galactose 4-epimerase (Gal10). The engineered yeast strain designated LN1, coexpressing all these enzymes, achieved an LNnT production level of 1.54 g/L. To further enhance LNnT yield, the flux through the pathway was improved by employing both protein fusion and modular assembly strategies. The assembly of LgtA and LgtB using short peptide tags resulted in a significant increase in LNnT titers, reaching 2.42 g/L. Finally, the LNnT titer reached 6.25 g/L under fed-batch fermentation conditions in a 500 mL flask, representing the highest reported production level of LNnT in S. cerevisiae to date.
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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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