Bioprospecting Fungal Glycosyl Hydrolases: Multi-locus Phylogenetic Analysis and Enzyme Activity Profiling for Enhanced Biomass Valorization

Abstract Lignocelluloses comprise of celluloses and hemicelluloses which can be effectively depolymerized to obtain fermentable sugars using diverse microbial enzymes, for subsequent conversion to various value-added products. Present study reports the bioprospecting of industrially significant microorganisms and their characterization to attain xylanases with high catalytic efficiency. Four potential xylanolytic fungi were identified through distinct primary and secondary screening process of 294 isolates from samples containing plant degrades. Morphological characterization and multigene analysis (ITS rDNA, 18S rDNA, nLSU rDNA, β-tubulin and actin gene) confirmed them Aspergillus niger AUMS56, Aspergillus tubingensis AUMS60, Aspergillus niger AUMS64 and Aspergillus fumigatus AUKEMS24 and their crude xylanase activities through submerged fermentation using corncob were 18.9, 32.29, 30.68 and 15.82 U ml-1, respectively. AUMS60 and AUMS64 have highest catalytic activity of 1429 U g-1 and 1243 U g-1, respectively, all having pH and temperature optima of 6.0 and 60°C respectively, where AUMS60 produced single xylanase (Xyn60; 36 kDa) and AUMS64 secreted 2 probable isozymes (Xyn64A and Xyn64B; 33.4 and 19.8 kDa). Maximum saccharification efficiency of AUMS60 and AUMS64 were 51.1% (13 h) and 52.2% (24 h) showing enhanced catalytic activity with various cations. Present research reports potential xylanases from indigenous fungi, providing opportunity for development of bio-catalysts concoction (novelty established) for enhanced saccharification of complex lignocelluloses finding specific industrial applications for production of value-added components.

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OPTIMIZATION AND CHARACTERIZATION OF EXO-POLYGALACTURONASE BY Aspergillus niger CULTURED VIA SOLID STATE FERMENTATION

Jurnal Teknologi ◽

10.11113/jt.v81.12222 ◽

2018 ◽

Vol 81 (1) ◽

Author(s):

Halifah Pagarra ◽

Roshanida A. Rahman ◽

Nur Izyan Wan Azelee ◽

Rosli Md Illias

Keyword(s):

Solid State ◽

Aspergillus Niger ◽

Moisture Content ◽

Solid State Fermentation ◽

Incubation Time ◽

Industrial Applications ◽

Screening Process ◽

Industrial Sectors ◽

Polygalacturonase Production

Polygalacturonases represent an important member of pectinases group of enzymes with immense industrial applications. The activity of exo-polygalacturonase produced by Aspergillus niger was studied in solid state fermentation (SSF) using Nephrolepis biserrata leaves as substrate. Central composite design (CCD) was used to optimize four significant variables resulted from the screening process that has been initially analyzed for the production of exo-polygalacturonase which are incubation time, temperature, concentration of pectin and moisture content. The optimum exo-polygalacturonase production obtained was 54.64 U/g at 120 hours of incubation time, temperature at 340C, 5.0 g/L of pectin concentration and 75.26% of moisture content. For partial characterization of exo-polygalacturonase, the optimum temperature and pH were obtained at 50°C and pH 4.0, respectively. SDS-PAGE analysis showed that molecular weight of exo-polygalacturonase were 35 and 71 kDa. This study has revealed a significant production of exo-polygalacturonase by A. niger under SSF using cheap and easily available substrate and thus could found immense potential application in industrial sectors and biotechnology

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Low-Cost Cellulase-Hemicellulase Mixture Secreted by Trichoderma harzianum EM0925 with Complete Saccharification Efficacy of Lignocellulose

International Journal of Molecular Sciences ◽

10.3390/ijms21020371 ◽

2020 ◽

Vol 21 (2) ◽

pp. 371 ◽

Cited By ~ 7

Author(s):

Yu Zhang ◽

Jinshui Yang ◽

Lijin Luo ◽

Entao Wang ◽

Ruonan Wang ◽

...

Keyword(s):

Lignocellulosic Biomass ◽

Trichoderma Harzianum ◽

Low Cost ◽

Value Added ◽

Industrial Applications ◽

Fermentable Sugars ◽

Enzyme Preparations ◽

Ultrafine Grinding ◽

Saccharification Efficiency ◽

Important Intermediate

Fermentable sugars are important intermediate products in the conversion of lignocellulosic biomass to biofuels and other value-added bio-products. The main bottlenecks limiting the production of fermentable sugars from lignocellulosic biomass are the high cost and the low saccharification efficiency of degradation enzymes. Herein, we report the secretome of Trichoderma harzianum EM0925 under induction of lignocellulose. Numerously and quantitatively balanced cellulases and hemicellulases, especially high levels of glycosidases, could be secreted by T. harzianum EM0925. Compared with the commercial enzyme preparations, the T. harzianum EM0925 enzyme cocktail presented significantly higher lignocellulolytic enzyme activities and hydrolysis efficiency against lignocellulosic biomass. Moreover, 100% yields of glucose and xylose were obtained simultaneously from ultrafine grinding and alkali pretreated corn stover. These findings demonstrate a natural cellulases and hemicellulases mixture for complete conversion of biomass polysaccharide, suggesting T. harzianum EM0925 enzymes have great potential for industrial applications.

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A novel approach of methane dehydroaromatization using group VIB metals (Cr, Mo, W) supported on sulfated zirconia

MRS Advances ◽

10.1557/adv.2020.368 ◽

2020 ◽

Vol 5 (63) ◽

pp. 3407-3417

Author(s):

Md Ashraful Abedin ◽

Swarom Kanitkar ◽

James J. Spivey

Keyword(s):

Catalytic Activity ◽

Sulfated Zirconia ◽

Value Added ◽

Product Yield ◽

Industrial Applications ◽

Acid Sites ◽

Single Step ◽

Intermediate Step ◽

Similar Reaction ◽

Methane Dehydroaromatization

AbstractMethane dehydroaromatization (MDHA) is a direct activation approach to covert methane to value-added chemicals in a single step. This requires no intermediate step, making it a commercially economic approach. Mo supported on HZSM-5/MCM-22 is a well-studied catalyst for this reaction, where Mo sites are responsible for activating methane to C2Hy dimers, which can oligomerize on HZSM-5 Bronsted acid sites to produce aromatics. Challenges for these bifunctional catalysts involve rapid coking and low product yield. In this study, a novel catalytic approach is introduced using group VIB metals (Cr, Mo, W) supported on sulfated zirconia (SZ) solid acid. It is believed that the Bronsted acidity of SZ should help to convert the dimers generated from metal sites to ethylene and aromatics like benzene.Here, fresh Mo, W and Cr were doped into SZ and characterized using pyridine DRIFTS, ammonia TPD, BET and SEM-EDS.. Catalytic activity for MDHA was ranked as Mo>W>Cr. Mo/SZ showed greater selectivity towards ethylene and benzene, followed by W/SZ, which was selective primarily towards ethylene. Cr/SZ showed the least activity under similar reaction conditions, producing only a small amount of ethylene. Higher catalytic activity for Mo/SZ was possibly due to reduced Mo oxide sites, found from XANES analysis, as well as higher acidity, observed from TPD. Deactivation was mainly due to coking, observed from subsequent TPO analysis. Further investigation is necessary to enhance the activity of this novel catalytic approach before considering for potential industrial applications.

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Rational Engineering of a Cold-Adapted α-Amylase from the Antarctic Ciliate Euplotes focardii for Simultaneous Improvement of Thermostability and Catalytic Activity

Applied and Environmental Microbiology ◽

10.1128/aem.00449-17 ◽

2017 ◽

Vol 83 (13) ◽

Cited By ~ 13

Author(s):

Guang Yang ◽

Hua Yao ◽

Matteo Mozzicafreddo ◽

Patrizia Ballarini ◽

Sandra Pucciarelli ◽

...

Keyword(s):

Catalytic Activity ◽

Catalytic Efficiency ◽

Industrial Applications ◽

Enzyme Engineering ◽

Content Type ◽

Cold Adapted ◽

Wide Range ◽

Cold Active ◽

Surface Loops ◽

The Antarctic

ABSTRACT The α-amylases are endo-acting enzymes that hydrolyze starch by randomly cleaving the 1,4-α-d-glucosidic linkages between the adjacent glucose units in a linear amylose chain. They have significant advantages in a wide range of applications, particularly in the food industry. The eukaryotic α-amylase isolated from the Antarctic ciliated protozoon Euplotes focardii (EfAmy) is an alkaline enzyme, different from most of the α-amylases characterized so far. Furthermore, EfAmy has the characteristics of a psychrophilic α-amylase, such as the highest hydrolytic activity at a low temperature and high thermolability, which is the major drawback of cold-active enzymes in industrial applications. In this work, we applied site-directed mutagenesis combined with rational design to generate a cold-active EfAmy with improved thermostability and catalytic efficiency at low temperatures. We engineered two EfAmy mutants. In one mutant, we introduced Pro residues on the A and B domains in surface loops. In the second mutant, we changed Val residues to Thr close to the catalytic site. The aim of these substitutions was to rigidify the molecular structure of the enzyme. Furthermore, we also analyzed mutants containing these combined substitutions. Biochemical enzymatic assays of engineered versions of EfAmy revealed that the combination of mutations at the surface loops increased the thermostability and catalytic efficiency of the enzyme. The possible mechanisms responsible for the changes in the biochemical properties are discussed by analyzing the three-dimensional structural model. IMPORTANCE Cold-adapted enzymes have high specific activity at low and moderate temperatures, a property that can be extremely useful in various applications as it implies a reduction in energy consumption during the catalyzed reaction. However, the concurrent high thermolability of cold-adapted enzymes often limits their applications in industrial processes. The α-amylase from the psychrophilic Antarctic ciliate Euplotes focardii (named EfAmy) is a cold-adapted enzyme with optimal catalytic activity in an alkaline environment. These unique features distinguish it from most α-amylases characterized so far. In this work, we engineered a novel EfAmy with improved thermostability, substrate binding affinity, and catalytic efficiency to various extents, without impacting its pH preference. These characteristics can be considered important properties for use in the food, detergent, and textile industries and in other industrial applications. The enzyme engineering strategy developed in this study may also provide useful knowledge for future optimization of molecules to be used in particular industrial applications.

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Effects of a Twin-Screw Extruder Equipped with a Molten Resin Reservoir on the Mechanical Properties and Microstructure of Recycled Waste Plastic Polyethylene Pellet Moldings

Polymers ◽

10.3390/polym13071058 ◽

2021 ◽

Vol 13 (7) ◽

pp. 1058

Author(s):

Hikaru Okubo ◽

Haruka Kaneyasu ◽

Tetsuya Kimura ◽

Patchiya Phanthong ◽

Shigeru Yao

Keyword(s):

Mechanical Properties ◽

Mechanical Performance ◽

Value Added ◽

Industrial Applications ◽

Twin Screw Extruder ◽

Screw Extruder ◽

Polymer Properties ◽

Wide Range ◽

Twin Screw ◽

Recycled Plastics

Each year, increasing amounts of plastic waste are generated, causing environmental pollution and resource loss. Recycling is a solution, but recycled plastics often have inferior mechanical properties to virgin plastics. However, studies have shown that holding polymers in the melt state before extrusion can restore the mechanical properties; thus, we propose a twin-screw extruder with a molten resin reservoir (MSR), a cavity between the screw zone and twin-screw extruder discharge, which retains molten polymer after mixing in the twin-screw zone, thus influencing the polymer properties. Re-extruded recycled polyethylene (RPE) pellets were produced, and the tensile properties and microstructure of virgin polyethylene (PE), unextruded RPE, and re-extruded RPE moldings prepared with and without the MSR were evaluated. Crucially, the elongation at break of the MSR-extruded RPE molding was seven times higher than that of the original RPE molding, and the Young’s modulus of the MSR-extruded RPE molding was comparable to that of the virgin PE molding. Both the MSR-extruded RPE and virgin PE moldings contained similar striped lamellae. Thus, MSR re-extrusion improved the mechanical performance of recycled polymers by optimizing the microstructure. The use of MSRs will facilitate the reuse of waste plastics as value-added materials having a wide range of industrial applications.

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Optimization of β-1,4-Endoxylanase Production by an Aspergillus niger Strain Growing on Wheat Straw and Application in Xylooligosaccharides Production

Molecules ◽

10.3390/molecules26092527 ◽

2021 ◽

Vol 26 (9) ◽

pp. 2527

Author(s):

Zahra Azzouz ◽

Azzeddine Bettache ◽

Nawel Boucherba ◽

Alicia Prieto ◽

Maria Jesus Martinez ◽

...

Keyword(s):

Aspergillus Niger ◽

Wheat Straw ◽

Plant Biomass ◽

Residual Activity ◽

Exchange Chromatography ◽

Glycosyl Hydrolases ◽

Biomass Degradation ◽

Xylan Hydrolysis ◽

Rsm Optimization ◽

Single Carbon Source

Plant biomass constitutes the main source of renewable carbon on the planet. Its valorization has traditionally been focused on the use of cellulose, although hemicellulose is the second most abundant group of polysaccharides on Earth. The main enzymes involved in plant biomass degradation are glycosyl hydrolases, and filamentous fungi are good producers of these enzymes. In this study, a new strain of Aspergillus niger was used for hemicellulase production under solid-state fermentation using wheat straw as single-carbon source. Physicochemical parameters for the production of an endoxylanase were optimized by using a One-Factor-at-a-Time (OFAT) approach and response surface methodology (RSM). Maximum xylanase yield after RSM optimization was increased 3-fold, and 1.41- fold purification was achieved after ultrafiltration and ion-exchange chromatography, with about 6.2% yield. The highest activity of the purified xylanase was observed at 50 °C and pH 6. The enzyme displayed high thermal and pH stability, with more than 90% residual activity between pH 3.0–9.0 and between 30–40 °C, after 24 h of incubation, with half-lives of 30 min at 50 and 60 °C. The enzyme was mostly active against wheat arabinoxylan, and its kinetic parameters were analyzed (Km = 26.06 mg·mL−1 and Vmax = 5.647 U·mg−1). Wheat straw xylan hydrolysis with the purified β-1,4 endoxylanase showed that it was able to release xylooligosaccharides, making it suitable for different applications in food technology.

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Enhancing the thermostability and activity of uronate dehydrogenase from Agrobacterium tumefaciens LBA4404 by semi-rational engineering

Bioresources and Bioprocessing ◽

10.1186/s40643-019-0267-3 ◽

2019 ◽

Vol 6 (1) ◽

Cited By ~ 1

Author(s):

Hui-Hui Su ◽

Fei Peng ◽

Pei Xu ◽

Xiao-Ling Wu ◽

Min-Hua Zong ◽

...

Keyword(s):

Agrobacterium Tumefaciens ◽

Rational Design ◽

Glucuronic Acid ◽

Specific Activity ◽

Value Added ◽

Catalytic Efficiency ◽

Site Directed Mutagenesis ◽

Glucaric Acid ◽

Triple Mutant ◽

Pharmaceutical Industries

Abstract Background Glucaric acid, one of the aldaric acids, has been declared a “top value-added chemical from biomass”, and is especially important in the food and pharmaceutical industries. Biocatalytic production of glucaric acid from glucuronic acid is more environmentally friendly, efficient and economical than chemical synthesis. Uronate dehydrogenases (UDHs) are the key enzymes for the preparation of glucaric acid in this way, but the poor thermostability and low activity of UDH limit its industrial application. Therefore, improving the thermostability and activity of UDH, for example by semi-rational design, is a major research goal. Results In the present work, three UDHs were obtained from different Agrobacterium tumefaciens strains. The three UDHs have an approximate molecular weight of 32 kDa and all contain typically conserved UDH motifs. All three UDHs showed optimal activity within a pH range of 6.0–8.5 and at a temperature of 30 °C, but the UDH from A. tumefaciens (At) LBA4404 had a better catalytic efficiency than the other two UDHs (800 vs 600 and 530 s−1 mM−1). To further boost the catalytic performance of the UDH from AtLBA4404, site-directed mutagenesis based on semi-rational design was carried out. An A39P/H99Y/H234K triple mutant showed a 400-fold improvement in half-life at 59 °C, a 5 °C improvement in $$ {\text{T}}_{ 5 0}^{ 1 0} $$ T 50 10 value and a 2.5-fold improvement in specific activity at 30 °C compared to wild-type UDH. Conclusions In this study, we successfully obtained a triple mutant (A39P/H99Y/H234K) with simultaneously enhanced activity and thermostability, which provides a novel alternative for the industrial production of glucaric acid from glucuronic acid.

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Promoting inherent catalytic activity and stability of TiO2 supported Pt single-atoms at CeOx-TiO2 interfaces

Journal of Materials Chemistry A ◽

10.1039/d1ta08059h ◽

2021 ◽

Author(s):

Mi Yoo ◽

Eunji Kang ◽

Hyuk Choi ◽

Hyunwoo Ha ◽

Han Seul Choi ◽

...

Keyword(s):

Catalytic Activity ◽

Catalytic Efficiency ◽

Reaction Centers ◽

Next Generation ◽

Single Atoms ◽

General Concern

Single-atoms Single-atoms (SAs) with atomically coordinated reaction centers are considered the next generation catalyst that can reveal exceptional catalytic efficiency. However, the general concern about thermodynamic vulnerabilities of the SAs...

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PHOTO-CATALYTIC ACTIVITY AND PHOTO-ABSORPTION OF PLASMA-SPRAYED NANO-STRUCTURED TiO2 COATINGS

International Journal of Modern Physics B ◽

10.1142/s021797921005483x ◽

2010 ◽

Vol 24 (31) ◽

pp. 6115-6127 ◽

Cited By ~ 1

Author(s):

MARYAMOSSADAT BOZORGTABAR ◽

MEHDI SALEHI ◽

MOHAMMADREZA RAHIMIPOUR ◽

MOHAMMADREZA JAFARPOUR

Keyword(s):

Catalytic Activity ◽

Flow Rate ◽

Test Chamber ◽

Catalytic Efficiency ◽

Surface Characteristics ◽

Catalytic Performance ◽

Atmospheric Plasma ◽

Argon Flow Rate ◽

Photo Catalytic Activity ◽

Argon Flow

Titanium dioxide coatings were deposited by utilizing atmospheric plasma-spraying system. The agglomerated P25/20 nano-powder and different spraying parameters (e.g., Argon flow rate and spray distance) were used to determine their influences on the microstructure, crystalline structure, photo-absorption, and photo-catalytic performance of the coatings. The microstructure and phases of as-sprayed TiO 2 coatings were characterized by scanning electron microscope SEM and X-ray diffraction, respectively. Surface characteristics were investigated by Fourier Transform Infrared. Photo-catalytic efficiency of the elaborated samples was also determined in an environmental test chamber set-up and evaluated from the conversion rate of ethanol. The photo-absorption was determined by UV–Vis spectrophotometer. The as-sprayed TiO2 coating was photo-catalytically reactive for the degradation of ethanol. The photo-catalytic activity was influenced by spray conditions. It is found that the photo-catalytic activity is significantly influenced by anatase content, surface area, and surface state. The results showed that the argon flow rate has an influence on the microstructure, anatase content, and photo-catalytic activity of the TiO 2 coatings.

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Dynamism and Cooperativity Essential for Efficient Catalysis in Aspergillus Niger Monoamine Oxidase

10.26434/chemrxiv.7707857.v1 ◽

2019 ◽

Author(s):

Christian Curado-Carballada ◽

Ferran Feixas ◽

Sílvia Osuna

Keyword(s):

Aspergillus Niger ◽

Monoamine Oxidase ◽

Active Site ◽

Conformational Dynamics ◽

Catalytic Efficiency ◽

Building Blocks ◽

Chiral Amine ◽

Evolutionary Pathway ◽

Accelerated Molecular Dynamics ◽

Open States

Aspergillus niger Monoamine Oxidase (MAO-N) is a homodimeric enzyme responsible for the oxidation of amines into the corresponding imine. Laboratory evolved variants of MAO-N in combination with a non-selective chemical reductant represents a powerful strategy for the deracemisation of chiral amine mixtures and, thus, is of interest for obtaining chiral amine building blocks. MAO-N presents a rich conformational dynamics with a flexible ß-hairpin region that can adopt closed, partially closed and open states. Despite the ß-hairpin conformational dynamics is altered along the laboratory evolutionary pathway of MAO-N, the connection between the ß-hairpin conformational dynamics and active site catalysis still remains unclear. In this work, we use accelerated molecular dynamics to elucidate the potential interplay between the ß-hairpin conformational dynamics and catalytic activity in MAO-N wild type and its evolved D5 variant. Our study reveals a delicate communication between both MAO-N subunits that impacts the active site architecture, and thus its catalytic efficiency. In both MAO-N WT and the laboratory evolved D5 variant, the ß-hairpin conformation in one of the monomers affects the productive binding of the substrate in the active site of the other subunit. However, both MAO-N WT and D5 variants show a quite different behaviour due to the distal mutations introduced experimentally with Directed Evolution.

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