Native Aspergillus Niger Research Articles

Physicochemical parameters optimization and peroxidase characterization from Aspergillus niger native strain by solid-state fermentation for improved dye decolorization

The hyper yield of peroxidase (POX) was investigated for a novel native Aspergillus niger strain identified by 18S RNA analysis. A. niger strains sequences were submitted to GenBank; IDs allotted were MN611114.1 (BMB 17) and MN559756.1 (BMB-18). The identified Aspergillus strains in combination showed enhanced (POX) activity (601.5 U/mL) by solid-state fermentation in comparison to their individual activities. POX was purified by ammonium sulfate, and size exclusion gel chromatography exhibited a 7.83-fold increase in POX concentration (13.3 U/mg) in comparison to BMB17 and BMB 18 (11.8 & 7.6 U/mg respectively). The best POX activity was obtained with pH 6.5, 37 °C, and 5 days of incubation. Using guaiacol as substrate, POX showed maximum activity (Vmax) of 537 U/mL with a corresponding Michaelis constant (Km) value of 126 µM. Calcium chloride worked as a POX activator at 300 & 400 mM. Zinc sulfate (500 mM), EDTA (5 mM); ethanol, propanol, and acetonitrile (50%) inhibited (18-30%) POX. Urea (1M), and copper sulfate (500 mM) strongly inhibited POX up to 40%. Polysorbate-80 (1%) slightly reduced the POX by 10% to 15%. BMB17+18-induced promising dye decolorization (88-98%) against all vat dyes, methylene blue, and phenol red.

Preliminary studies into transmogrification of Aspergillus genome for over expression of cellulase

The aim of this investigation was achieved by exposing native Aspergillus niger (WldAN) to an ultraviolet (UV) germicidal lamp (256 nm) kept at a distance of 40 cm for a time interval ranging from 2 –14 minutes, in a total of 56 minutes and finally using the improved strain to synthesize a more stable cellulase. The results showed growth improvement of the modified isolate with a density of 3.54 g/ml at the 9th hour as compared to 3.36 g/ml density of the native strain at the 7th hour on the 3rd day. Cellulase produced by submerged fermentation at 25 oC for 4 days using the native Aspergillus niger (WldAn) and modulated Aspergillus niger (UVAn) respectively recorded activities of 3.50 U/ml at 60 oC and 2.85 U/ml at pH 4.0 as compared to 2.40 U/ml at 50 oC and 2.24 U/ml at pH 5.0 recorded for cellulase synthase from the native strain

Understanding Thermostability Factors of Aspergillus niger PhyA Phytase: A Molecular Dynamics Study

Molecular dynamics simulation was used to study the dynamic differences between native Aspergillus niger PhyA phytase and a mutant with 20% greater thermostability. Atomic root mean square deviation, radius of gyration, and number of hydrogen bonds and salt bridges are examined to determine thermostability factors. The results suggest that, among secondary structure elements, loops have the most impact on the thermal stability of A. niger phytase. In addition, the location rather than the number of hydrogen bonds is found to have an important contribution to thermostability. The results also show that salt bridges may have stabilizing or destabilizing effect on the enzyme and influence its thermostability accordingly.

Xylanolytic enzymes production by Aspergillus niger GS1 from solid-state fermentation on corn stover and their effect on ruminal digestibility

Hemicellulosic agricultural by-products such as corn stover (CS) are highly available materials which represent an opportunity to develop value added products. Native Aspergillus niger GS1 was used for solid-state fermentation (SSF) on alkali pre-treated CS (ACS) aimed to optimize xylanolytic enzymes production, and their effect on in vitro ruminal and true digestibility of ACS. Enzyme production was empirically modelled using a fractional factorial design 2 9-5 , and the resulting significant factors were glucose, yeast extract and two mineral salts, which were arranged in a Draper-Lin optimization design at two levels. Predicted optimum xylanolytic activity of 33.6 U (mg protein) -1 was achieved at 48 hrs of SSF, and was validated by confirmatory experiments. ACS was incubated with a semipurified enzymatic extract (EE) showing a xylanolytic activity of 1600 U kg -1 dry ACS for 12 hrs before exposure to cow's ruminal liquid for 72 hrs, which led to 5% and 10% increase of in vitro ruminal and true digestibility, respectively. CS is a readily available by-product in different regions which after alkaline treatment and partial hydrolysis with the EE, may be advantageously used as supplement for ruminant feed.

Homologue expression of a β-xylosidase from native Aspergillus niger

Xylan constitutes the second most abundant source of renewable organic carbon on earth and is located in the cell walls of hardwood and softwood plants in the form of hemicellulose. Based on its availability, there is a growing interest in production of xylanolytic enzymes for industrial applications. β-1,4-xylan xylosidase (EC 3.2.1.37) hydrolyses from the nonreducing end of xylooligosaccharides arising from endo-1,4-β-xylanase activity. This work reports the partial characterization of a purified β-xylosidase from the native strain Aspergillus niger GS1 expressed by means of a fungal system. A gene encoding β-xylosidase, xlnD, was successfully cloned from a native A. niger GS1 strain. The recombinant enzyme was expressed in A. niger AB4.1 under control of A. nidulans gpdA promoter and trpC terminator. β-xylosidase was purified by affinity chromatography, with an apparent molecular weight of 90 kDa, and showed a maximum activity of 4,280 U mg protein(-1) at 70°C, pH 3.6. Half-life was 74 min at 70°C, activation energy was 58.9 kJ mol(-1), and at 50°C optimum stability was shown at pH 4.0-5.0. β-xylosidase kept residual activity >83% in the presence of dithiothreitol (DTT), β-mercaptoethanol, sodium dodecyl sulfate (SDS), ethylenediaminetetraacetate (EDTA), and Zn(2+). Production of a hemicellulolytic free xylosidase showed some advantages in applications, such as animal feed, enzymatic synthesis, and the fruit-juice industry where the presence of certain compounds, high temperatures, and acid media is unavoidable in the juice-making process.

Optimization of enantioselective resolution of racemic ibuprofen by native lipase from Aspergillus niger

Resolution of (R,S)-ibuprofen (2-(4-isobutylphenyl)propionic acid) enantiomers by esterification reaction with 1-propanol in different organic solvents was studied using native Aspergillus niger lipase. The main variables controlling the process (enzyme concentration and 1-propanol:ibuprofen molar ratio) have been optimized using response surface methodology based on a five-level, two-variable central composite rotatable design, in which the selected objective function was enantioselectivity. This enzyme preparation showed preferentially catalyzes the esterification of R(-)-ibuprofen, and under optimum conditions (7% w/v of enzyme and molar ratio of 2.41:1) the enantiomeric excess of active S(+)-ibuprofen and total conversion values were 79.1 and 48.0%, respectively, and the E-value was 32, after 168 h of reaction in isooctane.

Identification and elimination by site-directed mutagenesis of thermolabile aspartyl bonds in Aspergillus awamori glucoamylase.

Both native Aspergillus niger glucoamylase and wild-type Aspergillus awamori glucoamylase expressed in Saccharomyces cerevisiae, which have identical primary structures, undergo hydrolysis at aspartyl bonds at low pH values and elevated temperatures. In native A.niger enzyme the Asp126-Gly127 bond was preferentially cleaved at pH 3.5, while at pH 4.5 cleavage of the Asp257-Pro258 and Asp293-Gly294 bonds was dominant. In wild-type A.awamori glucoamylase, cleavage of the latter was dominant at both pH 3.5 and 4.5. Site-directed mutations Asp126-->Glu and Gly127-->Ala in wild-type enzyme decreased specific activities by approximately 60 and 30%, respectively, and increased irreversible thermoinactivation rates 3- to 4-fold at pH 4.5. Replacement of Asp257 with Glu and Asp293 with Glu or Gln decreased specific activities by approximately 20%, but greatly reduced cleavage of the Asp257-Pro258 and Asp293-Gly294 bonds. The Asp257-->Glu mutant was produced very slowly and was more thermostable than wild-type glucoamylase at pH 4.5 up to 70 degrees C. Replacement of Asp293 with either Glu or Gln significantly raised protein production and slightly increased thermostability at pH 3.5 and 4.5, but not at pH 5.5.

Fermentation of a yeast producing A. niger glucose oxidase: scale-up, purification and characterization of the recombinant enzyme.

We have developed a fermentation process to produce up to 3 grams per liter of active, secreted glucose oxidase from a recombinant Saccharomyces cerevisiae. Real-time size-exclusion HPLC analysis is used to monitor enzyme production during fermentation, and purification to more than 95 percent is obtained using only filtration methods. The recombinant enzyme is stable to higher temperatures and a wider pH range than the native Aspergillus niger enzyme, and is free of contaminating amylase, cellulase and catalase.