The chimeric GaaR-XlnR transcription factor induces pectinolytic activities in the presence of D-xylose in Aspergillus niger
| Autor: | Kun, Roland S., Garrigues, Sandra, Di Falco, Marcos, Tsang, Adrian, de Vries, Ronald P., Sub Molecular Plant Physiology, Molecular Plant Physiology |
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| Přispěvatelé: | Sub Molecular Plant Physiology, Molecular Plant Physiology, Westerdijk Fungal Biodiversity Institute - Fungal Physiology, Westerdijk Fungal Biodiversity Institute |
| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
0106 biological sciences
Aspergillus niger/genetics Mutant Transcription Factors/genetics 01 natural sciences Applied Microbiology and Biotechnology Fungal Proteins 03 medical and health sciences D-xylose 010608 biotechnology Gene Expression Regulation Fungal CRISPR Inducer Transcription factor Gene CRISPR/Cas9 030304 developmental biology Applied Genetics and Molecular Biotechnology chemistry.chemical_classification 0303 health sciences Xylose biology Cas9 Aspergillus niger General Medicine Pectinases biology.organism_classification Chimeric transcription factor Fungal Proteins/genetics Enzyme Fungal Biochemistry chemistry Gene Expression Regulation Transcription Factors Biotechnology |
| Zdroj: | Applied Microbiology and Biotechnology, 105(13), 5553. Springer Verlag Applied Microbiology and Biotechnology Applied Microbiology and Biotechnology, 105(13), 5553-5564. Springer Verlag GmbH |
| ISSN: | 0175-7598 |
| Popis: | Abstract Aspergillus niger is a filamentous fungus well known for its ability to produce a wide variety of pectinolytic enzymes, which have many applications in the industry. The transcriptional activator GaaR is induced by 2-keto-3-deoxy-L-galactonate, a compound derived from D-galacturonic acid, and plays a major role in the regulation of pectinolytic genes. The requirement for inducer molecules can be a limiting factor for the production of enzymes. Therefore, the generation of chimeric transcription factors able to activate the expression of pectinolytic genes by using underutilized agricultural residues would be highly valuable for industrial applications. In this study, we used the CRISPR/Cas9 system to generate three chimeric GaaR-XlnR transcription factors expressed by the xlnR promoter by swapping the N-terminal region of the xylanolytic regulator XlnR to that of the GaaR in A. niger. As a test case, we constructed a PpgaX-hph reporter strain to evaluate the alteration of transcription factor specificity in the chimeric mutants. Our results showed that the chimeric GaaR-XlnR transcription factor was induced in the presence of D-xylose. Additionally, we generated a constitutively active GaaR-XlnR V756F version of the most efficient chimeric transcription factor to better assess its activity. Proteomics analysis confirmed the production of several pectinolytic enzymes by ΔgaaR mutants carrying the chimeric transcription factor. This correlates with the improved release of D-galacturonic acid from pectin by the GaaR-XlnR V756F mutant, as well as by the increased L-arabinose release from the pectin side chains by both chimeric mutants under inducing condition, which is required for efficient degradation of pectin. Key points • Chimeric transcription factors were generated by on-site mutations using CRISPR/Cas9. • PpgaX-hph reporter strain allowed for the screening of functional GaaR-XlnR mutants. • Chimeric GaaR-XlnR induced pectinolytic activities in the presence of D-xylose. |
| Databáze: | OpenAIRE |
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