Abstract
Background: Synthetic L-ascorbic acid (vitamin C) is
widely used as a preservative and nutrient in food and
pharmaceutical industries. In the current production
method, D-glucose is converted to L-ascorbic acid via
several biochemical and chemical steps. The main source
of L-ascorbic acid in human nutrition is plants. Several
alternative metabolic pathways for L-ascorbic acid
biosynthesis are known in plants. In one of them,
D-galacturonic acid is the precursor. D-Galacturonic acid
is also the main monomer in pectin, a plant cell wall
polysaccharide. Pectin is abundant in biomass and is
readily available from several waste streams from fruit
and sugar processing industries.
Results: In the present work, we engineered the
filamentous fungus Aspergillus niger for the conversion
of D-galacturonic acid to L-ascorbic acid. In the
generated pathway, the native D-galacturonate reductase
activity was utilized while the gene coding for the
second enzyme in the fungal D-galacturonic acid pathway,
an L-galactonate consuming dehydratase, was deleted. Two
heterologous genes coding for enzymes from the plant
L-ascorbic acid pathway - L-galactono-1,4-lactone
lactonase from Euglena gracilis (EgALase) and
L-galactono-1,4-lactone dehydrogenase from Malpighia
glabra (MgGALDH) - were introduced into the A. niger
strain. Alternatively, an unspecific L-gulono-1,4-lactone
lactonase (smp30) from the animal L-ascorbic acid pathway
was introduced in the fungal strain instead of the plant
L-galactono-1,4-lactone lactonase. In addition, a strain
with the production pathway inducible with D-galacturonic
acid was generated by using a bidirectional and
D-galacturonic acid inducible promoter from the fungus.
Even though, the lactonase enzyme activity was not
observed in the resulting strains, they were capable of
producing L-ascorbic acid from pure D-galacturonic acid
or pectin-rich biomass in a consolidated bioprocess.
Product titers up to 170 mg/l were achieved.
Conclusions: In the current study, an L-ascorbic acid
pathway using D-galacturonic acid as a precursor was
introduced to a microorganism for the first time. This is
also the first report on an engineered filamentous fungus
for L-ascorbic acid production and a proof-of-concept of
consolidated bioprocess for the production.
Original language | English |
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Article number | 2 |
Number of pages | 9 |
Journal | Microbial Cell Factories |
Volume | 14 |
DOIs | |
Publication status | Published - 2015 |
MoE publication type | A1 Journal article-refereed |
Keywords
- L-ascorbic acid
- D-galacturonic acid
- L-galactonic acid
- Citrus peel
- Aspergillus niger
- Metabolic engineering