Comparative evaluation of Aspergillus niger strains for endogenous pectin-depolymerization capacity and suitability for d-galacturonic acid production

Schäfer, Dominik; Schmitz, Kevin; Weuster‐Botz, Dirk; Benz, J. Philipp

doi:10.1007/s00449-020-02347-z

Cited by 15 publications

(21 citation statements)

References 55 publications

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“…This analysis also included a comparison between N400 and N402. The results indicated that the growth profile in bioreactor was similar, although the N402 strain reached a higher maximum dry biomass level compared to N400 (2.28 ± 0.12 g/L for N400 vs 3.38 ± 0.07 g/L for N402) (Schäfer et al, 2020). The morphology of the mycelium of the N400 and N402 strains during cultivation on pectin was also examined and compared.…”

Section: Discussionmentioning

confidence: 97%

“…An important consideration is to which extent the mutations in the kinesin-like protein and the transcription factor that lead to shorter conidiophore stalk and reduced radial growth identified in this study might affect the conclusions reached in previous studies. Schäfer et al (2020) performed a comparative study in which different A. niger strains were compared for growth under controlled conditions. This analysis also included a comparison between N400 and N402.…”

Section: Discussionmentioning

confidence: 99%

“…The total pectinase and endo-specific polygalacturonase activity of medium samples was also analyzed. Compared to N400, the N402 strain had both reduced total pectinase and endo-polygalacturonase activity (Schäfer et al, 2020). A possible role for the cspA or cspB mutations in regulating pectinase expression was also suggested by the reduced expression of plyA [pectate lyase A and pelA (pectin lyase A)] in N400 compared to N402 derivatives (Parenicova, 2000).…”

Section: Discussionmentioning

confidence: 99%

“…Studies using A. niger as a model investigating transcriptional regulation of (biopolymer degrading) enzyme production are also numerous (e.g., van Peij et al, 1998;Yuan et al, 2008;de Souza et al, 2011;Delmas et al, 2012;Kowalczyk et al, 2015;Alazi et al, 2016). Based on the observation that the expression of pectinases might be altered in the N402 strain (Parenicova, 2000;Schäfer et al, 2020), mutations in N402 might also have an effect on the expression of these enzymes network. To circumvent possible effects of the mutations introduced in N402, N400 should be considered as a true wild-type strain and used preferably.…”

Section: Discussionmentioning

confidence: 99%

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Genetic Characterization of Mutations Related to Conidiophore Stalk Length Development in Aspergillus niger Laboratory Strain N402

et al. 2021

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Aspergillus niger is an important filamentous fungus in industrial biotechnology for the production of citric acid and enzymes. In the late 1980s, the A. niger N400/NRRL3 strain was selected for both fundamental and applied studies in relation to several processes including gluconic acid and protein production. To facilitate handling of A. niger, the N400 wild-type strain was UV mutagenized in two consecutive rounds to generate N401 and N402. N402 was used as a reference laboratory strain and exhibits the phenotypes with reduced conidiophore stalk length and reduced radial growth. The conidiophore stalk length and radial growth of A. niger strain N400 were determined and compared to N401 and N402. The length of N400 conidiophore stalks (2.52 ± 0.40 mm) was reduced in N401 and N402 to 0.66 ± 0.14 mm and 0.34 ± 0.06 mm, respectively. Whereas N400 reached a colony diameter of 6.7 ± 0.2 cm after 7 days, N401 and N402 displayed reduced radial growth phenotype (4.3 ± 0.1 and 4.1 ± 0.1, respectively). To identify the mutations (dubbed cspA and cspB) responsible for the phenotypes of N401 and N402, the genomes were sequenced and compared to the N400 genome sequence. A parasexual cross was performed between N400 and N402 derivatives to isolate segregants which allowed cosegregation analysis of single nucleotide polymorphisms and insertions and deletions among the segregants. The shorter conidiophore stalk and reduced radial growth in N401 (cspA) was found to be caused by a 9-kb deletion on chromosome III and was further narrowed down to a truncation of NRRL3_03857 which encodes a kinesin-like protein homologous to the A. nidulans UncA protein. The mutation responsible for the further shortening of conidiophore stalks in N402 (cspB) was found to be caused by a missense mutation on chromosome V in a hitherto unstudied C2H2 transcription factor encoded by the gene NRRL3_06646. The importance of these two genes in relation to conidiophore stalk length and radial growth was confirmed by single and double gene deletion studies. The mutations in the laboratory strain N402 should be taken into consideration when studying phenotypes in the N402 background.

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Section: Discussionmentioning

confidence: 97%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Genetic Characterization of Mutations Related to Conidiophore Stalk Length Development in Aspergillus niger Laboratory Strain N402

et al. 2021

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“…No growth, no sorbitol consumption, no D-GalA consumption, and no L-GalOA production were observed with S. cerevisiae SiHY001 (data not shown). Enzymatic hydrolysis of SBPP had been performed in 100 mM sodium acetate buffer (Schäfer et al 2020). As acetic acid is known to affect the metabolism of S. cerevisiae already at concentrations as low as 3 mM (Sousa et al 2012), growth of S. cerevisiae SiHY001 was studied in miniaturized single-use stirred tank reactors with diverse buffers and buffer concentrations at pH 5.0 in SC medium (sodium acetate, sodium citrate, potassium acetate, and potassium citrate).…”

Section: Growth Of S Cerevisiae Sihy001 With Varying Buffer Componentsmentioning

confidence: 99%

D-Galacturonic acid reduction by S. cerevisiae for L-galactonate production from extracted sugar beet press pulp hydrolysate

Wagner

Schäfer

Haimerl

et al. 2021

Appl Microbiol Biotechnol

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Pectin-rich residues are considered as promising feedstocks for sustainable production of platform chemicals. Enzymatic hydrolysis of extracted sugar beet press pulp (SBPP) releases the main constituent of pectin, d-galacturonic acid (d-GalA). Using engineered Saccharomyces cerevisiae, d-GalA is then reduced to l-galactonate (l-GalOA) with sorbitol as co-substrate. The current work addresses the combination of enzymatic hydrolysis of pectin in SBPP with a consecutive optimized biotransformation of the released d-GalA to l-GalOA in simple batch processes in stirred-tank bioreactors. Process conditions were first identified with synthetic media, where a product concentration of 9.9 g L-1 L-GalOA was obtained with a product selectivity of 99% (L-GalOA D-GalA-1) at pH 5 with 4% (w/v) sorbitol within 48 h. A very similar batch process performance with a product selectivity of 97% was achieved with potassium citrate buffered SBPP hydrolysate, demonstrating for the first time direct production of L-GalOA from hydrolyzed biomass using engineered S. cerevisiae. Combining the hydrolysis process of extracted SBPP and the biotransformation process with engineered S. cerevisiae paves the way towards repurposing pectin-rich residues as substrates for value-added chemicals. Key points • Efficient bioreduction of D-GalA with S. cerevisiae in stirred-tank reactors • Batch production of L-GalOA by engineered S. cerevisiae with high selectivity • Direct L-GalOA production from hydrolyzed sugar beet press pulp Graphical abstract

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Optimizing pectin lyase production using the one‐factor‐at‐a‐time method and response surface methodology

Gül,

Dursun,

Tepe

et al. 2024

Biotech and App Biochem

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Pectinases are commonly industrially synthesized by molds. This study aimed to optimize pectin lyase synthesis by a bacterium, Pseudomonas fluorescens, using both the one‐factor‐at‐a‐time (OFAT) method and response surface methodology. First, on optimization of pectin lyase fermentation by the OFAT method, the effects of pectin, peptone, yeast extract, (NH4)2SO4, pH, and salts were investigated. The highest pectin lyase activity was determined to be 28.63 U/mL at pH 8, 30°C, with 1% (w/v) pectin and 0.14% (w/v) (NH4)2SO4 concentration at the 90th hour. The effect of substrate inhibition on the microbial growth was also investigated, and the results showed that the process can be described by noncompetitive inhibition model. The values of kinetic constants were determined as µm = 0.175 h−1, KS = 6.931 g/L, and, KI = 6.932 g/L by nonlinear regression analysis. It was reported that pectin lyase enzymes exhibited peak activity at 50°C and pH 8. Finally, response surface methodology (RSM) was utilized to optimize pH, concentrations of ammonium sulfate, and pectin, which were chosen as independent variables. The interactions between these variables were also examined. According to RSM, the optimum values of the parameters to achieve a maximum pectin lyase activity of 35.62 U/mL were determined to be pH 7.97, 1.25% (w/v) pectin concentration, and 0.25% (w/v) (NH4)2SO4 concentration.

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Comparative evaluation of Aspergillus niger strains for endogenous pectin-depolymerization capacity and suitability for d-galacturonic acid production

Cited by 15 publications

References 55 publications

Genetic Characterization of Mutations Related to Conidiophore Stalk Length Development in Aspergillus niger Laboratory Strain N402

Genetic Characterization of Mutations Related to Conidiophore Stalk Length Development in Aspergillus niger Laboratory Strain N402

D-Galacturonic acid reduction by S. cerevisiae for L-galactonate production from extracted sugar beet press pulp hydrolysate

Optimizing pectin lyase production using the one‐factor‐at‐a‐time method and response surface methodology

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