Cooked off-flavor was produced during the processing of concentrated peach puree (CPP), which led to aroma deterioration. Enzymatic treatment was beneficial in eliminating off-flavors and improving the aroma quality. Herein, the efficacy of glycosidase (AR2000), glucose oxidation (GOD), and their combination on the inhibition of off-flavors and aroma enhancement were evaluated. Compared with CPP, contents of benzaldehyde, benzyl alcohol, nonanal, and linalool increased by 198%, 1222%, 781%, and 71% after AR2000 treatment via the metabolisms of shikimate, glucose, linoleic acid, and linolenic acid, leading to the strengthening of floral and grassy. Due to the removal of 1-octen-3-one via linolenic acid metabolism, cooked off-flavor could be significantly weakened by GOD. Furthermore, Furthermore, the combination of AR2000 and GOD could not only inhibit the production of 1-octen-3-one to weaken the cooked note but also enhance grassy and floral attributes via the increase of aldehydes and alcohols.

Protein kinase C substrate 80K-H (PRKCSH) plays a crucial role in the protein N-terminal glycosylation process, with emerging evidence implicating its involvement in tumorigenesis. To comprehensively assess PRKCSH\'s significance across cancers, we conducted a pan-cancer analysis using data from The Cancer Genome Atlas (TCGA), Genotype-Tissue Expression (GTEx), and Cancer Cell Line Encyclopedia (CCLE). We assessed aberrant PRKCSH mRNA and protein expression, examined its prognostic implications, and identified correlations with clinical features, tumor mutational burden (TMB), microsatellite instability (MSI), and tumor immunity across cancer types. We explored PRKCSH gene alterations, DNA methylation, and their impact on patient prognosis. Gene Set Enrichment Analysis (GSEA) and single-cell analysis revealed potential biological roles. Additionally, we investigated drug susceptibility and conducted Connectivity Map (Cmap) analysis. Key findings revealed that PRKCSH exhibited overexpression in most tumors, with a significant association with poor overall survival (OS) in six cancer types. Notably, PRKCSH expression demonstrated variations across disease stages, primarily increasing in advanced stages among eleven tumor types. Moreover, PRKCSH exhibited significant correlations with TMB in five cancer categories, MSI in eight, and displayed associations with immune cell populations in pan-cancer analysis. Genetic variations in PRKCSH were identified across 26 tumor types, suggesting favorable disease-free survival. Furthermore, PRKCSH methylation displayed a significant negative correlation with its expression in 27 tumor types, with a marked decrease compared to normal tissues in ten tumors. Cmap predicted 24 potential therapeutic small molecules in over four cancer types. This study highlights that PRKCSH, as a potential oncogene, may be a promising prognostic marker and therapeutic target of immunotherapy for a range of malignancies.

Although pervasive microplastics (MPs) pollution in terrestrial ecosystems invites increasing global concern, impact of MPs on soil microbial community assembly and ecosystem multifunctionality received relatively little attention. Here, we manipulated a mesocosm experiment to investigate how polyethylene MPs (PE MPs; 0, 1%, and 5%, w/w) influence ecosystem functions including plant production, soil quality, microbial community diversity and assembly, enzyme activities in carbon (C), nitrogen (N) and phosphorus (P) cycling, and multifunctionality in the maize-soil continuum. Results showed that PE MPs exerted negligible effect on plant biomass (dry weight). The treatment of 5% PE MPs caused declines in the availability of soil water, C and P, whereas enhanced soil pH and C storage. The activity of C-cycling enzymes (α/β-1, 4-glucosidase and β-D-cellobiohydrolase) was promoted by 1% PE MPs, while that of β-1, 4-glucosidase was inhibited by 5% PE MPs. The 5% PE MPs reduced the activity of N-cycling enzymes (protease and urease), whereas increased that of the P-cycling enzyme (alkaline phosphatase). The 5% PE MPs shifted soil microbial community composition, and increased the number of specialist species, microbial community stability and networks resistance. Moreover, PE MPs altered microbial community assembly, with 5% treatment decreasing dispersal limitation proportion (from 13.66% to 9.96%). Overall, ecosystem multifunctionality was improved by 1% concentration, while reduced by 5% concentration of PE MPs. The activity of α/β-1, 4-glucosidase, urease and protease, and ammonium-N content were the most important predictors of ecosystem multifunctionality. These results underscore that PE MPs can alter soil microbial community assembly and ecosystem multifunctionality, and thus development and implementation of practicable solutions to control soil MPs pollution become increasingly imperative in sustainable agricultural production.

The in-vitro digestive properties of myofibrillar protein (MP) in mirror carp (Cyprinus carpio L.) after freeze-thaw (F-T) cycles were analyzed in terms of the relationship between protein degradation, oxidation, and structural properties. The F-T samples exhibited a significant increase in glucosidase activity, N-acetyl-β-d-glucosidase activity, total protease activity, and non-protein nitrogen content. α-aminoadipate semialdehyde and γ-glutamate semialdehyde contents increased by 23.17% and 123.12%, respectively. Furthermore, 53.97% decrease in the total nitrogen content and changes in the content of different soluble proteins were observed. X-ray diffraction intensity, thermal stability, free amine content, hydrolysis degree, and digestibility of the MP samples decreased, and the 2θ angle and zeta potential were reversed. Besides, changes in the amide band wavenumbers were also detected. Therefore, the protein structure was unfolded and aggregates were formed through degradation and oxidation induced by the F-T cycles, ultimately making the in-vitro digestion of MP difficult.

Hyperthermophilic Sulfolobus solfataricus β-glycosidase (SS-βGly), with higher stability and activity than mesophilic enzymes, has potential for industrial ginsenosides biotransformation. However, its relatively low ginsenoside Rd-hydrolyzing activity limits the production of pharmaceutically active minor ginsenoside compound K (CK). In this study, first, we used molecular docking to predict the key enzyme residues that may hypothetically interact with ginsenoside Rd. Then, based on sequence alignment and alanine scanning mutagenesis approach, key variant sites were identified that might improve the enzyme catalytic efficiency. The enzyme catalytic efficiency (kcat/Km) and substrate affinity (Km) of the N264D variant enzyme for ginsenoside Rd increased by 60% and decreased by 17.9% compared with WT enzyme, respectively, which may be due to a decrease in the binding free energy (∆G) between the variant enzyme and substrate Rd. In addition, Markov state models (MSM) analysis during the whole 1000-ns MD simulations indicated that altering N264 to D made the variant enzyme achieve a more stable SS-βGly conformational state than the wild-type (WT) enzyme and corresponding Rd complex. Under identical conditions, the relative activities and the CK conversion rates of the N264D enzyme were 1.7 and 1.9 folds higher than those of the WT enzyme. This study identified an excellent hyperthermophilic β-glycosidase candidate for industrial biotransformation of ginsenosides.

In this paper, a novel bifunctional cellulase gene cel1 was cloned from Thermoascus aurantiacus by PCR and heterologously expressed in Pichia pastoris GS115. Bioinformatics and other related tools were used to compare the nucleotide homology of target genes, and analyze the signal peptide, transmembrane domain, hydrophilicity, secondary and tertiary structure of proteins. It was concluded that cel1 has similar endoglucanase nucleotide sequences and falls under the GH5 family. It was also found that cel1 has nucleotide sequences similar to glucosidase, which can infer that cel1 may have the properties of glucosidase, indicating that cel1 is multifunctional. At the same time, a part of the nucleotide sequence of the gene was removed to obtain a new gene cel2, and after highly efficient heterologous expression, its specific activity was found to be 2.1 times higher. Its enhancement is related to the exposure of the protein\'s hollow three-dimensional structure. This paper provides good material for exploring the relationship between the structure of bifunctional enzymes and their functions, which lays a solid foundation for further research and applications, and provides useful insight for gene mining of other novel enzymes.

Ormosia hosiei Hemsl. et Wils. is an economical and medicinal plant, increasingly cultivated in China; however, its branches and leaves are often pruned as waste. This is the first study focused on the phytochemical profiles and antioxidant, anti-α-glucosidase, anti-tyrosinase, and anti-neuroinflammatory activities of the branches and leaves of O. hosiei. Herein, thirty-seven characteristic compounds were identified by UPLC-MS/MS and twelve were detected for the first time in O. hosiei. Twenty-seven phenolics were further quantified and significant differences in phenolic compositions between the branches and leaves of O. hosiei were observed. The ethanol extracts exhibited promising antioxidant, anti-α-glucosidase, anti-tyrosinase, and anti-neuroinflammatory effects, and the bioactivities significantly correlated with total phenolic content and twelve individual phenolics. Naringin, genistein, vitexin, vitexin-2-O-rhamnoside, syringaresinol and syringaresinol-4-O-β-D-glucopyranoside can be considered potential quality markers of O. hosiei. Our results provided solid evidence that the branches and leaves of O. hosiei deserve more attention and exploitation, considering the potential to be developed as functional foods or herbal medicines.

In the barley β-D-glucan glucohydrolase, a glycoside hydrolase family 3 (GH3) enzyme, the Trp286/Trp434 clamp ensures β-D-glucosides binding, which is fundamental for substrate hydrolysis during plant growth and development. We employ mutagenesis, high-resolution X-ray crystallography, and multi-scale molecular modelling methods to examine the binding and conformational behaviour of isomeric β-D-glucosides during substrate-product assisted processive catalysis that operates in GH3 hydrolases. Enzyme kinetics reveals that the W434H mutant retains broad specificity, while W434A behaves as a strict (1,3)-β-D-glucosidase. Investigations of reactant movements on the nanoscale reveal that processivity is sensitive to mutation-specific alterations of the tryptophan clamp. While wild-type and W434H utilise a lateral cavity for glucose displacement and sliding of (1,3)-linked hydrolytic products through the catalytic site without dissociation, consistent with their high hydrolytic rates, W434A does not adopt processive catalysis. Phylogenomic analyses of GH3 hydrolases disclose the evolutionary advantage of the tryptophan clamp that confers broad specificity, high catalytic efficiency, and processivity.

Salicylic acid (SA) is a stress hormone synthesized in phenylalanine ammonia-lyase (PAL) and the branching acid pathway. SA has two interconvertible forms in plants: SAG (SA O-β-glucoside) and SA (free form). The molecular mechanism of conversion of SA to SAG had been reported previously. However, which genes regulate SAG to SA remained unknown. Here, we report a cytoplasmic β-glucosidase (β-Glu) which participates in the SA pathway and is involved in the brown hull pigmentation in rice grain. In the current study, an EMS-generated mutant brown hull 1 (bh1) displayed decreased contents of SA in hulls, a lower photosynthesis rate, and high-temperature sensitivity compared to the wild type (WT). A plaque-like phenotype (brown pigmentation) was present on the hulls of bh1, which causes a significant decrease in the seed setting rate. Genetic analysis revealed a mutation in LOC_Os01g67220, which encodes a cytoplasmic Os1βGlu4. The knock-out lines displayed the phenotype of brown pigmentation on hulls and decreased seed setting rate comparable with bh1. Overexpression and complementation lines of Os1βGlu4 restored the phenotype of hulls and normal seed setting rate comparable with WT. Subcellular localization revealed that the protein of Os1βGlu4 was localized in the cytoplasm. In contrast to WT, bh1 could not hydrolyze SAG into SA in vivo. Together, our results revealed the novel role of Os1βGlu4 in the accumulation of flavonoids in hulls by regulating the level of free SA in the cellular pool.

The formation and development of biological soil crusts (biocrusts) potentially affect the cycles and stoichiometric characteristics of soil carbon (C), nitrogen (N), and phosphorus (P). However, it is still unclear how soil microbes adapt to such changes. In this study, we examined the effects of moss-dominated biocrusts coverage (0, 1%-20%, 20%-40%, 40%-60%, 60%-80%, and 80%-100%) on soil physicochemical properties, soil microbial biomass, and ectoenzyme activities [β-1, 4-glucosidase (BG), β-1, 4-N-acetyl glucosidase (NAG), acid phosphatase (AP)] in two soil layers (0-5 and 5-10 cm) in the Three Gorges Reservoir area, as well as the covariations of soil-microbe-ectoenzyme C:N:P stoichiometry. The results showed that biocrust development significantly increased soil clay content, water stable aggregates, soil C, N, P contents, and significantly decreased soil bulk density and sand content. Microbial biomass C, N, P and ectoenzyme activities were significantly increased with increasing biocrust coverage. Soil depth did not affect soil physicochemical properties and C:N:P, but significantly affected microbial biomass, ectoenzyme activities, BG:AP and NAG:AP. Soil C, N and P contents were significantly positively correlated with microbial biomass and ectoenzyme activities, negatively correlated with BG:NAG, while positively correlated with NAG:AP, but had no significant correlation with microbial biomass C:N:P. There was no significant correlation between soil-microbe and microbial-ectoenzyme C:N:P. BG:NAG:AP decreased gradually with the increase of C:N:P stoichiometric imbalance between microbe and soil. This study indicated that the microbial metabolism was co-limited by N and P and with stronger P limitation. Microbes could maintain homeostasis by adjusting their own biomass and ectoenzyme C:N:P to adapt to changes in soil ecological stoichiometry driven by biocrust development.
生物结皮的形成和发育显著影响土壤碳(C)、氮(N)、磷(P)循环及其化学计量特征,土壤微生物如何适应环境资源的化学计量变化仍不明确。本研究以三峡库区苔藓结皮为对象,分析结皮盖度(0、1%～20%、20%～40%、40%～60%、60%～80%和80%～100%)对土壤理化性质(0～5和5～10 cm土层)、微生物生物量和胞外酶活性[(β-1,4-葡萄糖苷酶(BG)、β-1,4-N-乙酰氨基葡萄糖苷酶(NAG)、酸性磷酸酶(AP)]的影响,探索土壤-微生物-胞外酶C∶N∶P化学计量特征间的协变性。结果表明: 生物结皮发育显著提高了土壤黏粒、水稳性团聚体和土壤C、N、P含量,显著降低了土壤容重和砂粒含量;微生物生物量C、N、P和胞外酶活性均随结皮盖度的增大而显著增加;土层深度对土壤理化性质及C∶N∶P均无显著影响,但显著影响微生物生物量、胞外酶活性及BG∶AP和NAG∶AP。相关分析显示,土壤C、N、P含量与微生物生物量和胞外酶活性呈显著正相关,与BG∶NAG呈显著负相关,与NAG∶AP呈显著正相关,但与微生物生物量C∶N∶P无显著相关性;土壤-微生物、微生物-胞外酶C∶N∶P相关性均不显著,BG∶NAG∶AP随着微生物与土壤间C∶N∶P化学计量不平衡性的增加而逐渐降低。表明微生物养分代谢同时受N和P的限制,且P的限制较强烈,微生物可以通过调整自身生物量以及胞外酶C∶N∶P适应生物结皮发育驱动的土壤化学计量变化,从而维持内稳态。.