Abstract
| Original language | English |
|---|---|
| Article number | 210 |
| Journal | Microbial Cell Factories |
| Volume | 15 |
| DOIs | |
| Publication status | Published - 2016 |
| MoE publication type | A1 Journal article-refereed |
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Keywords
- aspergillus niger
- metabolic engineering
- CRISPR
- pectin
- d-galacturonic acid
- galactaric acid
- mucic acid
- uronate dehydrogenase
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Engineering Aspergillus niger for galactaric acid production: elimination of galactaric acid catabolism by using RNA sequencing and CRISPR/Cas9. / Kuivanen, Joosu (Corresponding Author); Wang, Jasmin; Richard, Peter.
In: Microbial Cell Factories, Vol. 15, 210, 2016.Research output: Contribution to journal › Article › Scientific › peer-review
TY - JOUR
T1 - Engineering Aspergillus niger for galactaric acid production: elimination of galactaric acid catabolism by using RNA sequencing and CRISPR/Cas9
AU - Kuivanen, Joosu
AU - Wang, Jasmin
AU - Richard, Peter
PY - 2016
Y1 - 2016
N2 - Background: meso-Galactaric acid is a dicarboxylic acid that can be produced by the oxidation of d-galacturonic acid, the main constituent of pectin. Mould strains can be engineered to perform this oxidation by expressing the bacterial enzyme uronate dehydrogenase. In addition, the endogenous pathway for d-galacturonic acid catabolism has to be inactivated. The filamentous fungus Aspergillus niger would be a suitable strain for galactaric acid production since it is efficient in pectin hydrolysis, however, it is catabolizing the resulting galactaric acid via an unknown catabolic pathway. Results : In this study, a transcriptomics approach was used to identify genes involved in galactaric acid catabolism. Several genes were deleted using CRISPR/Cas9 together with in vitro synthesized sgRNA. As a result, galactaric acid catabolism was disrupted. An engineered A. niger strain combining the disrupted galactaric and d-galacturonic acid catabolism with an expression of a heterologous uronate dehydrogenase produced galactaric acid from d-galacturonic acid. The resulting strain was also converting pectin-rich biomass to galactaric acid in a consolidated bioprocess. Conclusions: In the present study, we demonstrated the use of CRISPR/Cas9 mediated gene deletion technology in A. niger in an metabolic engineering application. As a result, a strain for the efficient production of galactaric acid from d-galacturonic acid was generated. The present study highlights the usefulness of CRISPR/Cas9 technology in the metabolic engineering of filamentous fungi.
AB - Background: meso-Galactaric acid is a dicarboxylic acid that can be produced by the oxidation of d-galacturonic acid, the main constituent of pectin. Mould strains can be engineered to perform this oxidation by expressing the bacterial enzyme uronate dehydrogenase. In addition, the endogenous pathway for d-galacturonic acid catabolism has to be inactivated. The filamentous fungus Aspergillus niger would be a suitable strain for galactaric acid production since it is efficient in pectin hydrolysis, however, it is catabolizing the resulting galactaric acid via an unknown catabolic pathway. Results : In this study, a transcriptomics approach was used to identify genes involved in galactaric acid catabolism. Several genes were deleted using CRISPR/Cas9 together with in vitro synthesized sgRNA. As a result, galactaric acid catabolism was disrupted. An engineered A. niger strain combining the disrupted galactaric and d-galacturonic acid catabolism with an expression of a heterologous uronate dehydrogenase produced galactaric acid from d-galacturonic acid. The resulting strain was also converting pectin-rich biomass to galactaric acid in a consolidated bioprocess. Conclusions: In the present study, we demonstrated the use of CRISPR/Cas9 mediated gene deletion technology in A. niger in an metabolic engineering application. As a result, a strain for the efficient production of galactaric acid from d-galacturonic acid was generated. The present study highlights the usefulness of CRISPR/Cas9 technology in the metabolic engineering of filamentous fungi.
KW - aspergillus niger
KW - metabolic engineering
KW - CRISPR
KW - pectin
KW - d-galacturonic acid
KW - galactaric acid
KW - mucic acid
KW - uronate dehydrogenase
U2 - 10.1186/s12934-016-0613-5
DO - 10.1186/s12934-016-0613-5
M3 - Article
VL - 15
JO - Microbial Cell Factories
JF - Microbial Cell Factories
SN - 1475-2859
M1 - 210
ER -