Search Results (72)

Fruit yield and quality improvement are challenges for researchers and farmers. This study reveals that the main fruit traits of blueberry (Vaccinium ashei ‘Bluegem’) were significantly improved after hydrogen (H₂)-based irrigation, assessed by the increased single fruit weight (14.59 ± 6.66%) and fruit equatorial diameter (4.19 ± 2.39%), decreased titratable acidity, increased solid–acid and sugar–acid ratios. The enhancement of fruit quality was confirmed by the increased total volatiles, vitamin C contents, and antioxidant capacity. Using weighted protein co-expression network analysis (WPCNA), proteomic interrogation revealed that serine carboxypeptidase-like proteins I/II (SCPLI/II), ADP ribosylation factor 1/2 (ARF1/2), and UDP-glucosyltransferase 85A (UGT85A) might be functionally associated with the increased fruit weight and size driven by H₂. Reduced organic acid accumulation was caused by the regulation of the specific enzymes involved in sucrose metabolism (e.g., α-amylase, endoglucanase, β-glucosidase, etc.). H₂ regulation of fatty acid degradation (e.g., acyl CoA oxidase 1 (ACX1), acetyl CoA acyltransferase 1 (ACAA1), etc.) and phenylpropanoid metabolism were used to explain the improved fruit aroma and anthocyanin accumulation. Meanwhile, the upregulated heat shock protein 20/70 matched with the enhanced antioxidant activity. Together, this study provides a novel approach for yield and quality improvement in horticultural crops. Full article

Ultraviolet (UV)-induced mutagenesis is a cost-effective and straightforward technique for introducing random genetic variations without the use of chemical reagents or genetic engineering. It is commonly employed to enhance enzyme activity in industrial trains. In this study, Trichoderma sp. was exposed to UV radiation at varying distances (4, 9, and 13 cm) and durations (2, 4, 6, and 8 min) to induce mutations. The activities of endoglucanase (EG), β-glucosidase (BGL), and cellobiohydrolase (CBH) were assessed following treatment. The 4 cm exposure distance yielded the highest enhancement, with EG, BGL, and CBH activities increasing 1.5-, 1.3-, and 0.9-fold, respectively. When the distance was fixed at 4 cm, the optimal exposure time was identified as 4 min, yielding further enhancements of 1.9-, 1.6-, and 1.4-fold, respectively. The resulting mutant, designated Mut-4, was scaled up in a 10-L bioreactor to assess its industrial applicability. Mut-4 retained its enhanced performance, achieving 1.9-, 2.0-, and 1.4-fold enhancements in EG, BGL, and CBH activities, respectively, compared with the original strain. These findings indicate that combining UV-induced mutagenesis with basic screening is an effective strategy for enhancing cellulolytic enzyme production, representing a promising approach for lignocellulosic biomass conversion. Full article

The fungi belonging to the genus Pleurotus can be cultivated in different substrates and represent excellent producers of several extracellular enzymes. In this study, we analyzed eleven hydrolytic enzymes of the P. eryngii and P. ostreatus secretomes, which were collected at three different growth stages after 23 days (mycelial colonization of about 50% of the substrate), 34 days (100% colonization of the substrate) and 50 days (after the first flush). Mushrooms were axenically cultivated on the same substrate. The results demonstrate that proteases, lipases, amylases, α-glucosidase, cellulases (endoglucanase, β-cellobiohydrolase and β-glucosidase) and hemicellulase (xylosidase, glucuronidase, arabinosidase and mannosidase) activities were higher in the secretomes from P. eryngii than those from P. ostreatus. Time course analysis revealed for both species a similar enzymatic activity profile, in which in the early stages of mycelium development, both species use starch as the main carbon source. Protease and lipase activities increased and remained constant during the subsequent formation of fruiting bodies, whereas cellulase and hemicellulase activities decreased after the complete mycelial colonization of the substrate. The zymographic analysis suggested the presence in the secretomes of proteolytic activities belonging to different classes. In conclusion, both mushroom species released into the secretomes a broad spectrum of hydrolytic enzymes potentially useful in various biotechnological fields. Full article

The efficient degradation of lignocellulose is essential for valorizing agricultural waste and reducing environmental pollution. An efficient degradation process requires an enzyme cocktail capable of comprehensively deconstructing lignocellulosic components. In this study, the secretome of Coriolopsis trogii Mafic-2001 induced by rice straw was examined, and the enzymatic composition and reaction conditions of Coriolopsis trogii were optimized. Mafic-2001 secreted an enzyme cocktail that included ligninolytic enzymes, cellulases, and hemicellulases. However, the relative abundances of endoglucanase (EG) and β-glucosidase (βG) were only 64.37% and 10.69%, respectively, compared with the relative abundance of cellobiohydrolase, which indicated a critical bottleneck in degradation efficiency. To overcome this limitation, the recombinant enzymes rEG1 and rβG1 were expressed in Pichia pastoris X-33. A functionally enhanced enzyme cocktail (rEG1–rβG1–Mafic-2001 = 0.05:0.09:0.86) was developed via a mixture design to achieve a reducing sugar yield of 2.77 mg/mL from Chinese distillers’ grains (CDGs). Structural analyses revealed that the optimized enzyme cocktail disrupted the reticulated fiber architecture of CDGs and attenuated the characteristic Fourier-transform infrared spectroscopy peaks of lignin, cellulose, and hemicellulose. This study elucidates the synergistic lignocellulose deconstruction mechanism of Mafic-2001 and establishes a precision enzyme-supplementation strategy for efficient CDG bioconversion, providing a scalable platform for the valorization of lignocellulosic biomass. Full article

Fungal species secrete various enzymes and are considered the primary sources of industrially important cellulases. Cellulases are essential natural factors for cellulose degradation and have attracted significant interest for multiple applications. However, reducing the cost and enhancing cellulase production remains a significant challenge. Mutagenesis has opened a new window for enhancing enzyme secretion by modifying the organism’s genome. In this study, cellulases from Alternaria citri were produced and characterized, and the optimization for ideal fermentation conditions was performed for three types of cellulases (endoglucanase, exoglucanase, and β-glucosidase) by a wild-type (A. citri) and a mutant strain (A. citri 305). Ethyl methanesulfonate, a chemical mutagen, was used to enhance cellulase production by A. citri. The results demonstrate the improved cellulolytic ability of the mutant strain A. citri 305 utilizing lignocellulosic waste substances, particularly, orange-peel powder, wheat straw, sugarcane bagasse, and sawdust, making this study economically valuable. This evokes the potential for multi-dimensional applications in enzyme production, waste degradation, and biofuel generation. This study highlights that the activity of cellulases to hydrolyze various lignocellulosic substrates is enhanced after mutagenesis. Full article

The increasing generation of agro-industrial waste and its improper disposal have raised significant environmental concerns, highlighting the urgent need for sustainable alternatives which would repurpose these materials. In this context, enzymes such as endoglucanase play a critical role in degrading lignin–cellulose biomass by catalyzing the breakdown of β-1,4-glycosidic bonds in cellulose, thereby converting it into fermentable sugars with diverse industrial applications. This study aimed to investigate the production, purification, and characterization of an endoglucanase produced by the fungus Pleurotus djamor PLO13, using coconut fiber, sugarcane bagasse, wheat bran, and pineapple crown as substrates. Endoglucanase activity was measured by the Miller method (1959), using 2% (w/v) carboxymethyl cellulose (CMC) as substrate. Solid-state fermentation (SSF) was found to be highly efficient for enzyme synthesis, with wheat bran emerging as the most effective substrate, yielding an enzyme production of 7.19 U after 120 h of cultivation. The endoglucanase was purified through ethanol precipitation and ion-exchange chromatography using DEAE-Sepharose, achieving a recovery rate of 110%, possibly due to removal of inhibitors present in the crude extract. The purified enzyme exhibited stability across a broad pH range and thermostability, with optimal activity at pH 5.0 and 50 °C. Furthermore, the enzyme was activated by EDTA, Mn²⁺, and Ca²⁺, while being inhibited by Mg²⁺. Notably, the enzyme demonstrated halotolerance, with activity increasing by 60% upon the addition of 3 M NaCl. Kinetic analysis revealed that the purified enzyme showed affinity to the CMC substrate at the analyzed parameters (pH 5.0 and 50 °C), with Km and Vmax values of 0.0997 mg/mL and 112.2 µg/min/mL, respectively. These findings suggest that the endoglucanase from P. djamor PLO13 has promising potential for biotechnological applications, underscoring the feasibility of the use of lignocellulosic waste as sustainable substrates in industrial processes. Full article

The filamentous fungus Penicillium verruculosum (anamorph Talaromyces verruculosus) has been shown to be an efficient producer of secreted cellulases, used in biorefinery processes. Understanding the mechanisms of regulation of cellulase gene expression in the fungus P. verruculosum is a current task in industrial biotechnology, since it allows for targeted changes in the composition of the complex secreted by the fungus. Expression of cellulase genes in fungi is regulated mainly at the level of transcription via pathway-specific transcription factors (TF), the majority of which belong to the Zn(II)2Cys6 family of zinc binuclear cluster proteins. Transcriptional regulation of cellulase genes may have a species-specific pattern and involves several transcription factors. In this study, we used a qPCR method and transcriptome analysis to investigate the effect of knockouts and constitutive expression of genes encoding homologues of the regulatory factors XlnR and ClrB from P. verruculosum on the transcription of cbh1, egl2, and bgl1 genes, encoding three key cellulases, cellobiohydrolase, endoglucanase, and β-glucosidase, in the presence of various inducers. We have shown that the transcription factor XlnR of the filamentous fungus P. verruculosum is strictly responsible for the transcription of the main cellulolytic genes (cbh1, egl2, and bgl1) in the presence of xylose and xylobiose, but not in the presence of cellobiose. ClrB/Clr-2, a homologue from P. verruculosum, does not represent the main transcription factor regulating transcription of cellulolytic genes in the presence of selected inducers, unlike in the cases of Aspergillus nidulans, Aspergillus niger, and Penicillium oxalicum; apparently, it has a different function in fungi from the genus Talaromyces. We have also shown that constitutive expression of the transcription factor XlnR resulted in 3.5- and 2-fold increases in the activity of xylanase and β-glucosidase in a B1-XlnR enzyme preparation, respectively. In a practical sense, the obtained result can be used for the production of enzyme preparations based on the P. verruculosum B1-XlnR strain used for the bioconversion of renewable cellulose-containing raw materials into technical sugars. Full article

Seed shattering (SS) functions are a survival mechanism in plants, enabling them to withstand adverse environmental conditions and ensure reproduction. However, this trait limits seed yield. Psathyrostachys juncea, a perennial forage grass with many favorable traits, is constrained by SS, limiting its broader application. To investigate the mechanisms underlying SS, second-generation Illumina sequencing and third-generation PacBio sequencing were conducted on abscission zone tissues of P. juncea at 7, 14, 21, and 28 days after heading. GO enrichment analysis identified several significant biological processes, including the “cell wall macromolecule catabolic process”, “cell wall polysaccharide catabolic process”, “hemicellulose catabolic process”, and “xylan catabolic process”, all involved in cell wall degradation. KEGG enrichment analysis showed that differentially expressed genes were predominantly enriched in pathways related to “starch and sucrose metabolism”, “fructose and mannose metabolism”, “phenylpropanoid biosynthesis”, “pentose and glucuronate interconversions”, and “galactose metabolism”, each linked to both the synthesis and degradation of the cell wall. Further analysis of the “starch and sucrose metabolism” pathway revealed genes encoding fructokinase, hexokinase, β-glucosidase, sucrose phosphate synthase, sucrose synthase, and endoglucanase, all of which affected cellulose content. Reduced cellulose content can alter cell wall structure, leading to SS. These findings provide new insights into the regulation of SS in P. juncea and offer valuable references for other species within the Poaceae family. Full article

Pathogen-induced fruit decay is a significant threat to the kiwifruit industry, leading to considerable economic losses annually. The cell-wall-degrading enzymes (CWDEs) secreted by these pathogens are crucial for penetrating the cell wall and accessing nutrients. Among them, Diaporthe species are recognized as major causal agents of soft rot in kiwifruit, yet their pathogenic mechanisms are not well understood. In this study, we explored the production of various CWDEs secreted by Diaporthe Z1-1N, including polygalacturonase (PG), polymethylgalacturonase (PMG), polygalacturonic acid transeliminase (PGTE), pectin methyltranseliminase (PMTE), endoglucanase (Cx), and β-glucosidase (β-glu), both in liquid cultures and within infected kiwifruit tissues. Our findings revealed significant activities of two pectinases (PG and PMG) and cellulases (Cx and β-glu) in the infected tissues. In contrast, very low levels of PMTE and PGTE activities were observed under the same conditions. When orange pectin served as the carbon source, PG and PMG showed notable activities, while PMTE and PGTE remained inactive. Moreover, the activities of Cx and β-glu significantly decreased by more than 63 times in the liquid medium with carboxymethyl cellulose (CMC) as the carbon source compared to their levels in infected kiwifruit. A further analysis indicated that the necrotic lesions produced by pectinase extracts were larger than those produced by cellulase extracts. Notably, four enzymes—PG, PMG, Cx, and β-glu—exhibited high activities on the third or fourth day post-infection with Diaporthe Z1-1N. These results suggest that Diaporthe Z1-1N secretes a range of CWDEs that contribute to kiwifruit decay by enhancing the activities of PG, PMG, Cx, and β-glu. This study sheds light on the pathogenicity of Diaporthe in kiwifruit and highlights the importance of these enzymes in the decay process. Full article

To enhance the nutritional value of Acanthopanax senticosus leaves (AL), a fermentation process was conducted using a probiotic Bacillus mixture, and the changes in chemical constituents and biological activities before and after fermentation were compared. A response surface methodology was employed to optimize the liquid fermentation conditions of AL based on their influence on polyphenol content. Non-targeted metabolomics analysis was performed using LC-MS/MS to reveal the differing profiles of compounds before and after fermentation. The results indicated that Bacillus subtilis LK and Bacillus amyloliquefaciens M2 significantly influenced polyphenol content during fermentation. The optimal fermentation conditions were determined to be a fermentation time of 54 h, a temperature of 39.6 °C, and an inoculum size of 2.5% (v/v). In comparison to unfermented AL, the total polyphenol and flavonoid contents, as well as the free radical scavenging capacities measured by DPPH and ABTS assays, and the activities of β-glucosidase and endo-glucanase, were significantly increased. The non-targeted metabolomics analysis identified 1348 metabolites, of which 829 were classified as differential metabolites. A correlation analysis between the differential metabolites of polyphenols, flavonoids, and antioxidant activity revealed that 13 differential metabolites were positively correlated with antioxidant activity. Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis of the differential metabolites identified 82 pathways, with two of the top 25 metabolic pathways related to flavonoids. This study explores the potential for enhancing the active ingredients and biological effects of AL through probiotic fermentation using Bacillus strains. Full article

The objective of this study was to assess the cellulase production of four fungi: Aspergillus terreus NIH2624, Aspergillus clavatus NRRL1, Aspergillus versicolor CBS583.65 and Aspergillus phoenicis ATCC3157, under submerged cultivation conditions. When these fungi were cultured in shake flasks using Mandels and Weber’s minimal medium with 1% sugarcane bagasse as a carbon source and 1.8 g/L of rice bran extract as a nitrogen source, A. terreus showed maximum cellulase production (filter paper activity (FPase) 3.35 U/mL; carboxymethyl cellulase activity (CMCase) 1.69 U/mL). Consequently, A. terreus was selected for the optimization study for cellulase production. Among the different tested carbon sources, A. terreus showed higher CMCase activity when it was cultivated on delignified sugarcane bagasse (1.64 U/mL) and higher FPase activity on sugarcane straw (7.95 U/mL). Regarding the nitrogen sources, the maximum FPase activity was observed when using rice bran (FPase, 8.90 U/mL) and soybean meal (FPase, 9.63 U/mL). The optimized fermentation medium (minimal medium with delignified sugarcane bagasse and rice bran as carbon and nitrogen sources, respectively) resulted in an enzymatic cocktail mainly composed of xylanases, with a maximum activity of 1701.85 U/mL for beechwood xylan, 77.12 U/mL for endoglucanase and 21.02 U/mL for cellobiohydrolase. Additionally, the enzymatic cocktail showed efficient activities for β-glucosidase, β-xylanase, arabinofuranosidase and lytic polysaccharide monoxygenases (LPMOs). This cellulase enzyme solution has the potential to efficiently hydrolyze lignocellulosic biomass, producing second-generation sugars in biorefineries. Full article

Brown-rot fungi are large fungi that can decompose the cell walls of wood; they are notable for their secretion of diverse and complex enzymes that synergistically hydrolyze natural wood cellulose molecules. Fomitopsis pinicola (F. pinicola) is a brown-rot fungus of interest for its ability to break down the cellulose in wood efficiently. In this study, through a combination of rDNA-ITS analysis and morphological observation, the wood decay pathogen infecting Korean pine (Pinus koraiensis Siebold and Zucc.) was identified. Endoglucanase (CMCase) and β-glucosidase were quantified using the DNS (3,5-Dinitrosalicylic acid) method, and the cellulase activity was optimized using a single-factor method and orthogonal test. The results revealed that the wood-decaying fungus NE1 identified was Fomitopsis pinicola with the ITS accession number OQ880566.1. The highest cellulase activity of the strain reached 116.94 U/mL under the condition of an initial pH of 6.0, lactose 15 g·L⁻¹, KH₂PO₄ 0.5 g·L⁻¹, NH₄NO₃ 15 g·L⁻¹, MgSO₄ 0.5 g·L⁻¹, VB₁ 0.4 g·L⁻¹, inoculated two 5 mm fungal cakes in 80 mL medium volume cultured 28 °C for 5 days. This laid a foundation for improving the degradation rate of cellulose and biotransformation research, as well as exploring the degradation of cellulose by brown rot fungi. Full article

The enzyme-based degradation of lignocellulose for bioenergy production is an eco-friendly and sustainable approach. This study aimed to elucidate the enzymatic characteristics of endoglucanase (EGL), β-glucosidase (BGL), and xylanase (XYN) from Trichoderma guizhouence NJAU4742, and to explore the potential mechanisms underlying their synergistic degradation of different natural substrates. The results demonstrated that the three enzymes possessed remarkable high-temperature catalytic activity, broad pH adaptability, and responsiveness to different metal ions. The functional group absorption peaks of different substrates were shifted and altered after the synergistic action, particularly for C=O and O-H. Simultaneously, the crystallinity index of wheat straw, soybean straw, rice straw, and corn straw decreased by 7.40%, 2.37%, 20.60%, and 7.67%, respectively, compared to CK (natural straw). Additionally, the dense structure of different substrates was destroyed, and the inner parenchyma began to be exposed after the synergistic action, as observed by SEM. These findings offer valuable theoretical guidance for the development of lignocellulase applications. Full article

Cellulolytic microorganisms play a crucial role in agricultural waste disposal. Strain QXD-8^T was isolated from soil in northern China. Similarity analyses of the 16S rRNA gene, as well as the 120 conserved genes in the whole-genome sequence, indicate that it represents a novel species within the genus Microbacterium. The Microbacterium sp. QXD-8^T was able to grow on the CAM plate with sodium carboxymethyl cellulose as a carbon source at 15 °C, forming a transparent hydrolysis circle after Congo red staining, even though the optimal temperature for the growth and cellulose degradation of strain QXD-8^T was 28 °C. In the liquid medium, it effectively degraded cellulose and produced reducing sugars. Functional annotation revealed the presence of encoding genes for the GH5, GH6, and GH10 enzyme families with endoglucanase activity, as well as the GH1, GH3, GH39, and GH116 enzyme families with β-glucosidase activity. Additionally, two proteins in the GH6 family, one in the GH10, and two of nine proteins in the GH3 were predicted to contain a signal peptide and transmembrane region, suggesting their potential for extracellularly degrade cellulose. Based on the physiological features of the type strain QXD-8^T, we propose the name Microbacterium psychrotolerans for this novel species. This study expands the diversity of psychrotolerant cellulolytic bacteria and provides a potential microbial resource for straw returning in high-latitude areas at low temperatures. Full article

Interest in enzymes capable of neutralizing various mycotoxins is quite high. The methods used for the screening and selection of enzymes that catalyze the detoxification of mycotoxins should be sensitive and fast. However toxic compounds can be generated under the action of such enzymes. Thus, the assessment of the overall reduction in the toxic properties of reaction media towards bioluminescent bacteria seems to be the most reasonable control method allowing a quick search for the effective enzymatic biocatalysts. The influence of a wide range of mycotoxins and glucanases, which hydrolyze toxins with different chemical structures, on the analytical characteristics of luminescent photobacteria as a biosensing element has been studied. Different glucanases (β-glucosidase and endoglucanase) were initially selected for reactions with 10 mycotoxins based on the results of molecular docking which was performed in silico with 20 mycotoxins. Finally, the biorecognizing luminescent cells were used to estimate the residual toxicity of reaction media with mycotoxins after their interaction with enzymes. The notable non-catalytic decrease in toxicity of media containing deoxynivalenol was revealed with luminous cells for both types of tested glucanases, whereas β-glucosidase provided a significant catalytic detoxification of media with aflatoxin B₂ and zearalenone at pH 6.0. Full article