The hydrolysis of xylo-oligosaccharides catalyzed by β-xylosidase plays an important role in the degradation of lignocellulose. However, the enzyme is easily inhibited by its catalytic product xylose, which severely limits its application. Based on molecular docking, this paper studied the xylose affinity of Aspergillus niger β-xylosidase An-xyl, which was significantly differentially expressed in the fermentation medium of tea stalks, through cloning, expression and characterization. The synergistic degradation effect of this enzyme and cellulase on lignocellulose in tea stems was investigated. Molecular docking showed that the affinity of An-xyl to xylose was lower than that of Aspergillus oryzae β-xylosidase with poor xylose tolerance. The K_i value of xylose inhibition constant of recombinant-expressed An-xyl was 433.2 mmol/L, higher than that of most β-xylosidases of the GH3 family. The K_m and V_max towards pNPX were 3.6 mmol/L and 10 000 μmol/(min·mL), respectively. The optimum temperature of An-xyl was 65 ℃, the optimum pH was 4.0, 61% of the An-xyl activity could be retained upon treatment at 65 ℃ for 300 min, and 80% of the An-xyl activity could be retained upon treatment at pH 2.0-8.0 for 24 h. The hydrolysis of tea stem by An-xyl and cellulase produced 19.3% and 38.6% higher reducing sugar content at 2 h and 4 h, respectively, than that of using cellulase alone. This study showed that the An-xyl mined from differential expression exhibited high xylose tolerance and higher catalytic activity and stability, and could hydrolyze tea stem lignocellulose synergistically, which enriched the resource of β-xylosidase with high xylose tolerance, thus may facilitate the advanced experimental research and its application.
Aspergillus niger/genetics* ; Xylose/metabolism* ; Molecular Docking Simulation ; Xylosidases/genetics* ; Cellulases ; Tea ; Hydrogen-Ion Concentration ; Substrate Specificity

The L(+)-form of tartaric acid (L(+)-TA) exists extensively in nature, and is widely used in the food, chemical, textile, building, and pharmaceutical industries (Su et al., 2001). The main method for L(+)-TA production is microbial transformation by cis-epoxysuccinate hydrolase (CESH), which can catalyze the asymmetric hydrolysis of cis-epoxysuccinic acid or its salts to TA or tartrate (Bao et al., 2019). Seventeen species containing CESH have been isolated so far. However, most species for L(+)-TA production have been reported from bacteria (Xuan and Feng, 2019). The only fungus isolated from soil by our lab recently, that could be used as catalyst for the process under acidic condition, is Aspergillus niger WH-2 (Bao et al., 2020). In order to find strains with new characteristics, this study attempted to isolate a new CESH source from fungi and investigate its application value.
Aspergillus niger/metabolism* ; Biomass ; Catalysis ; Fermentation ; Hydrogen-Ion Concentration ; Hydrolases/chemistry* ; Hydrolysis ; Industrial Microbiology ; Magnetic Resonance Spectroscopy ; Penicillium/metabolism* ; Phylogeny ; Soil ; Species Specificity ; Stereoisomerism ; Tartrates/chemistry* ; Temperature ; Textiles

OBJECTIVE: A strain of Aspergillus niger (A. niger), capable of releasing bound phenolic acids from wheat bran, was isolated. This strain was identified by gene sequence identification. The antioxidant and anti-inflammatory capacity of ferulic acid released from wheat bran by this A. niger strain (FA-WB) were evaluated. METHODS: Molecular identification techniques based on PCR analysis of specific genomic sequences were conducted; antioxidant ability was examined using oxygen radical absorbance capacity (ORAC), cellular antioxidant activity (CAA) assays, and erythrocyte hemolysis assays. RAW264.7 cells were used as a model to detect anti-inflammatory activity. RESULTS: The filamentous fungal isolate was identified to be A. niger. ORAC and CAA assay showed that FA-WB had better antioxidant activity than that of the ferulic acid standard. The erythrocyte hemolysis assay results suggested that FA-WB could attenuate AAPH-induced oxidative stress through inhibition of reactive oxy gen species (ROS) generation. FA-WB could significantly restore the AAPH-induced increase in intracellular antioxidant enzyme activities to normal levels as well as inhibit the intracellular malondialdehyde formation. TNF-a, IL-6, and NO levels indicated that FA-WB can inhibit the inflammation induced by lipopolysaccharide (LPS). CONCLUSION Ferulic acid released from wheat bran by a new strain of A. niger had good anti-inflammatory activity and better antioxidant ability than standard ferulic acid.
Animals ; Anti-Inflammatory Agents ; metabolism ; pharmacology ; Antioxidants ; metabolism ; pharmacology ; Aspergillus niger ; genetics ; isolation & purification ; metabolism ; Coumaric Acids ; metabolism ; pharmacology ; DNA, Fungal ; analysis ; Dietary Fiber ; microbiology ; Erythrocytes ; drug effects ; metabolism ; Fermentation ; Hep G2 Cells ; Humans ; Interleukin-6 ; metabolism ; Lipopolysaccharides ; pharmacology ; Mice ; RAW 264.7 Cells ; Sheep ; Tumor Necrosis Factor-alpha ; metabolism

Carbon-limited continuous culture was used to study the relationship between the growth of Aspergillus niger and the production of glucoamylase. The result showed that when the specific growth rate was lower than 0.068 h(-1), the production of glucoamylase was growth-associated, when the specific growth rate was higher than 0.068 h(-1), the production of glucoamylase was not growth-associated. Based on the result of continuous culture, the Monod dynamics model of glucose consumption of A. niger was constructed, Combining Herbert-Pirt equation of glucose and oxygen consumption with Luedeking-Piret equation of enzyme production, the black-box model of Aspergillus niger for enzyme production was established. The exponential fed-batch culture was designed to control the specific growth rate at 0.05 h(-1) by using this model and the highest yield for glucoamylase production by A. niger reached 0.127 g glucoamylase/g glucose. The black-box model constructed in this study successfully described the glucoamylase production by A. niger and the result of the model fitted the measured value well. The black-box model could guide the design and optimization of glucoamylase production by A. niger.
Aspergillus niger ; metabolism ; Batch Cell Culture Techniques ; Carbon ; Culture Media ; Glucan 1,4-alpha-Glucosidase ; biosynthesis ; Glucose ; Industrial Microbiology ; methods ; Oxygen

The main commercial production of fructooligosaccharides (FOS) comes from enzymatic transformation using sucrose as substrate by microbial enzyme fructosyltransferase. A fructosyltransferase genomic DNA was isolated from Aspergillus niger QU10 by PCR. The nucleotide sequence showed a 1 941 bp size, and has been submitted to GenBank (KF699529). The cDNA of the fructosyltransferase, containing an open reading frame of 1 887 bp, was further cloned by RT-PCR. The fructosyltransferase gene from Aspergillus niger was functionally expressed both in Escherichia coli and Pichia pastoris GS 115. The highest activity value for the construction with the α-factor signal peptide reached 431 U/mL after 3 days of incubation. The recombinant enzyme is extensively glycosylated, and the active form is probably represented by a homodimer with an apparent molecular mass of 200 kDa as judged from mobility in seminative PAGE gels. The extracellular recombinant enzyme converted sucrose mostly to FOS, mainly 1-kestose and nystose, liberating glucose. FOS reached a maximal value and represented about 58% of total sugars present in the reaction mixture after 4 h reaction. The results suggest that the availability of recombinant Pichia pastoris as a new source of a FOS-producing enzyme might result of biotechnology interest for industrial application.
Aspergillus niger ; enzymology ; genetics ; Base Sequence ; Cloning, Molecular ; DNA, Complementary ; Escherichia coli ; Fungal Proteins ; genetics ; metabolism ; Glycosylation ; Hexosyltransferases ; genetics ; metabolism ; Molecular Sequence Data ; Molecular Weight ; Pichia ; Sucrose ; metabolism ; Trisaccharides ; metabolism

In order to discover the steroid biotransformation ability of filamentous fungus Aspergillus niger TCCC41650, we studied the fermentation of 4-androstene-3,17-dione with A. niger TCCC41650. The transformation product was purified, crystallized and determined as 16β-hydroxy-androst-4-ene-3,17-dione by X-ray single crystal diffraction method. The best fermentation condition was found to be pH 6.0, ethanol amount 2% with a substrate concentration of 1 per thousand, the transformation rate is 85.81% after 72 h. Based on the best of our knowledge, 16β-hydroxylation rarely occurs in microbial transformations of steroid. This study laid the foundation for the research of 16β-hydroxylation steroids
Androstenedione ; metabolism ; Aspergillus niger ; metabolism ; Biotransformation ; Fermentation ; Hydroxylation ; Industrial Microbiology

The monoamine oxidase mutant A-1 (F210V/L213C) from Aspergillus niger showed some catalytic activity on mexiletine. To futher improve its activity, the mutant was subjected to directed evolution with MegaWHOP PCR (Megaprimer PCR of Whole Plasmid) and selection employing a high-throughput agar plate-based colorimetric screen. This approach led to the identification of a mutant ep-1, which specific activity was 189% of that for A-1. The ep-1 also showed significantly improved enantioselectivity, with the E value increased from 101 to 282; its kinetic k(cat)/K(m) value increased from 0.001 51 mmol/(L x s) to 0.002 89 mmol/(L x s), suggesting that catalytic efficiency of ep-1 had been improved. The mutant showed obviously higher specific activities on 7 of all tested 11 amines substrates, and the others were comparable. Sequence analysis revealed that there was a new mutation T162A on ep-1. The molecular dynamics simulation indicated that T162A may affect the secondary structure of the substrate channel and expand the binding pocket.
Aspergillus niger ; enzymology ; Catalysis ; Kinetics ; Monoamine Oxidase ; genetics ; metabolism ; Mutation ; Polymerase Chain Reaction ; Protein Engineering ; Protein Structure, Secondary ; Substrate Specificity

In order to investigate the effects of fungal elicitor and macroporous adsorption resin on shikonin accumulation in hairy roots of arnebia euchroma (Royle) Johnst, we used spectrophotometry to determine the total naphthoquinone content of the hairy roots, by adding different volume ratio of Aspergillus niger elicitor, Aspergillus oryzae elicitor, and the macroporous resin into the M-9 liquid medium at different culture time. The results show that the total naphthoquinone content was 2.28 times higher than the control when we added mixed elicitors of Aspergillus niger and Aspergillus oryzae at the ratio of 2.5:50 in the 10th day of hairy roots cultivating. The total naphthoquinone content was 3.71 times higher than that of the control, when we added macroporous adsorption resin NKA-9. Aspergillus niger elicitor exhibited synergistic effect with Aspergillus oryzae elicitor to enhance the naphthoquinone. Also, the total naphthoquinone level was 4.17 times higher than that of the control by adding mixed fungal elicitor and macroporous adsorption resin NKA-9 in the bioreactor. Aspergillus oryzae and mixed elicitor could promote the hairy roots proliferation, and macroporous adsorption resin NKA-9 and mixed elicitor increased the total naphthoquinone content. In summary, the measure developed for Arnebia euchroma (Royle) Johnst hairy roots cultivating in bioreactors may potential for large-scale production of naphthoquinone.
Aspergillus niger ; metabolism ; Boraginaceae ; metabolism ; Fungal Proteins ; pharmacology ; Naphthoquinones ; analysis ; metabolism ; Plant Roots ; chemistry ; metabolism ; Porosity ; Resins, Synthetic ; pharmacology

Based on previous bioinformational analysis results, two Aspergillus niger lipase (ANL) mutants, ANL-Ser84Gly and ANL-Asp99Pro were constructed to screen ANL mutants with oil-water interface independence. ANL-Ser84Gly still displayed a pronounced interfacial activation, while ANL-Asp99Pro displayed no interfacial activation. The specific activity of ANL-Ser84Gly towards p-nitrophenyl palmitate (-myristate, -laurate and -decanoate) decreased by 29.8% (53.1, 60.1 and 77.1, respectively) than that of ANL, while the specific activity of ANL-Asp99Pro towards p-nitrophenyl palmitate increased by 2.2-fold. The mutation in the hinge region at both sides of the lid domain also destabilized various secondary structure factors of ANL-S84G and ANL-D99P, which resulted in a substantial decrease in thermostability. The achievement to construct oil-water interface-independent ANL mutants would help to further understand lipase interfacial activation mechanism.
Aspergillus niger ; enzymology ; genetics ; Base Sequence ; Enzyme Stability ; Fungal Proteins ; genetics ; metabolism ; Lipase ; genetics ; metabolism ; Molecular Sequence Data ; Mutant Proteins ; genetics ; Oils ; Substrate Specificity ; Water

Streptomyces S24 has broad spectrum against Aspergillus spp. in food and feed, such as Aspergillus flavus, Aspergillus niger and Asperegillus alutacells. The objective of this study was to improve the production of antifungal substances produced by S24 and to test their inhibitory effects on Aspergillus flavus. By using one-factor-at-a-time experiment and orthogonal design method, we optimized the fermentation medium. The composition of an optimized medium for the production of antifungal substances contained (g/L): starch soluble, 10; Glucose, 40; yeast extract, 8; soybean powder, 24; KH2PO4 4; and CaCO3 0.8. As a result, the productivity of antifungal substances could reach to 10 235.45 microg/mL, and this value was 2.81 times higher than that of initial medium before optimization. Additionally, inhibitory effects of the products on Aspergillus flavus were analyzed. Antagonistic tests indicated that the antifungal substances greatly inhibited mycelium growth and spores germination of Aspergillus flavus. We observed through microscope that the mycelia grew abnormally, such as contorting, bulging, vacuole increasing and the cytoplasmic contents inside effusing and the spores appeared unusual, such as gathering, deforming, cytoplasmic contents inside effusing and fracturing.
Adsorption ; Antifungal Agents ; chemistry ; isolation & purification ; pharmacology ; Aspergillus flavus ; drug effects ; growth & development ; Aspergillus niger ; drug effects ; growth & development ; Culture Media ; chemistry ; Food Contamination ; prevention & control ; Streptomyces ; chemistry ; growth & development ; metabolism