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Banana is the fourth most consumed crop worldwide, and its high economic value and health benefits have made it very popular. Bananas are climacteric fruits that ripen after harvesting. It has been reported that the endogenous substances in bananas change significantly during the ripening process. This study focused on levels of gamma-aminobutyric acid (GABA) and glutamic acid decarboxylase (GAD), an enzyme that catalyzes the synthesis of GABA, which reportedly fluctuates during the ripening stage. Previous studies have shown that GAD expression is associated with banana ripening; however, changes in its distribution during ripening have not been verified. This study aimed to clarify the relationship between GABA and GAD during ripening of ethylene-treated bananas. Visualization of the localization of endogenous GABA and GAD was performed using mass spectrometry imaging. To visualize GAD reaction, a glutamate-d 3 (labeled substrate) was supplied to the sample, and a GABA-d 3 (labeled product) was regarded as the localization of the enzymatic reaction. Liquid chromatography-mass spectrometry was also used to confirm the amount of GABA and activity of the GAD. This will allow us to clarify the direct relationship between GABA and GAD and to understand the role of the GAD reaction in phytohormones. [Display omitted] • Study clarifies GABA-GAD relationship during ripening of ethylene-treated bananas. • GABA and GAD levels change during banana ripening. • Study used mass spectrometry imaging for visualizing GABA amount and GAD reaction. • Liquid chromatography-mass spectrometry confirmed GABA amount and GAD reaction. [ABSTRACT FROM AUTHOR]

In recent years, many studies have focused on improving the bioconversion of cellulose by adding non-ionic surfactants. In our study, the effect of the addition of a polymer, polyethylene glycol (PEG 4000), on the bioconversion of different cellulose materials was evaluated, focusing on the hydrolysis efficiency and structural changes in pure cellulose after the enzymatic hydrolysis process. The obtained results showed that the addition of non-ionic surfactant significantly improved the digestibility of cellulosic materials. The highest hydrolysis efficiency was observed for Sigmacel 101 (Cel-S101) cellulose, which consists mainly of amorphous regions. In the case of Avicel cellulose (Cel-A), PEG had a lesser effect on the bioconversion's efficiency due to limited access to the crystal structure and limited substrate–cellulase interactions. The consistency of the obtained results is confirmed by qualitative and quantitative analyses (XRD, FTIR, and HPLC). Our findings may be helpful in further understanding the mechanism of the action of surfactants and improving the enzymatic hydrolysis process. [ABSTRACT FROM AUTHOR]

Abstract Enzyme reactions have numerous applications in diverse disciplines of science like chemistry, biology and biomechanics. In this study, we examine the role and act of enzymes in chemical reactions which is considered in the frame of fractional order model. The proposed model includes system of four equations which are studied via Caputo fractional operator. The systems of non-linear equations are evaluated by a semi-analytical approach called $$q$$ q -homotopy analysis transform method. The uniqueness and existence of the solutions has been investigated through fixed point theorem. The solutions of the proposed model are achieved through the considered method and the obtained outcomes are in the form of series which shows rapid convergence. The solutions are computed and graphs are plotted for the obtained results using mathematica software. The achieved results by the proposed method are unique and illustrate the significant dynamics of the considered model via 3D plots and graphs. The results of this study demonstrate the importance and effectiveness of projected derivative and technique in the analysis of time dependent fractional mathematical models. This study also gives an idea to extend the applications of enzymatic reactions in drug development, bio mechanics, and chemical reactions in various cellular metabolisms. Also, enzymatic reactions have a vital role in the fields of the food industry for processing food, in biotechnology for the manufacture of biofuels, and in metabolic engineering to design metabolic pathways.

Enzyme reactions have numerous applications in diverse disciplines of science like chemistry, biology and biomechanics. In this study, we examine the role and act of enzymes in chemical reactions which is considered in the frame of fractional order model. The proposed model includes system of four equations which are studied via Caputo fractional operator. The systems of non-linear equations are evaluated by a semi-analytical approach called q -homotopy analysis transform method. The uniqueness and existence of the solutions has been investigated through fixed point theorem. The solutions of the proposed model are achieved through the considered method and the obtained outcomes are in the form of series which shows rapid convergence. The solutions are computed and graphs are plotted for the obtained results using mathematica software. The achieved results by the proposed method are unique and illustrate the significant dynamics of the considered model via 3D plots and graphs. The results of this study demonstrate the importance and effectiveness of projected derivative and technique in the analysis of time dependent fractional mathematical models. This study also gives an idea to extend the applications of enzymatic reactions in drug development, bio mechanics, and chemical reactions in various cellular metabolisms. Also, enzymatic reactions have a vital role in the fields of the food industry for processing food, in biotechnology for the manufacture of biofuels, and in metabolic engineering to design metabolic pathways. [ABSTRACT FROM AUTHOR]

Enzymatic reaction kinetics are central in analyzing enzymatic reaction mechanisms and target-enzyme optimization, and thus in biomanufacturing and other industries. The enzyme turnover number (k cat) and Michaelis constant (K m), key kinetic parameters for measuring enzyme catalytic efficiency, are crucial for analyzing enzymatic reaction mechanisms and the directed evolution of target enzymes. Experimental determination of k cat and K m is costly in terms of time, labor, and cost. To consider the intrinsic connection between k cat and K m and further improve the prediction performance, we propose a universal pretrained multitask deep learning model, MPEK, to predict these parameters simultaneously while considering pH, temperature, and organismal information. Through testing on the same k cat and K m test datasets, MPEK demonstrated superior prediction performance over the previous models. Specifically, MPEK achieved the Pearson coefficient of 0.808 for predicting k cat, improving ca. 14.6% and 7.6% compared to the DLKcat and UniKP models, and it achieved the Pearson coefficient of 0.777 for predicting K m, improving ca. 34.9% and 53.3% compared to the Kroll_model and UniKP models. More importantly, MPEK was able to reveal enzyme promiscuity and was sensitive to slight changes in the mutant enzyme sequence. In addition, in three case studies, it was shown that MPEK has the potential for assisted enzyme mining and directed evolution. To facilitate in silico evaluation of enzyme catalytic efficiency, we have established a web server implementing this model, which can be accessed at http://mathtc.nscc-tj.cn/mpek. [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, the EFSA Panel on Nutrition, Novel Foods and Food Allergens (NDA) was asked to deliver an opinion on glucosyl hesperidin (GH) as a novel food (NF) pursuant to Regulation (EU) 2015/2283. The NF, which is produced from hesperidin and dextrin by enzymatic reactions, is a powder consisting mainly of monoglucosyl hesperidin (MGH) and unreacted hesperidin (flavonoid), which account in total for up to 92.8% (on dry basis) of the product. The applicant proposed to use the NF in specific drinks and food supplements leading to a maximum intake of up to 364 mg per day for adults. The target population is the general population, except for food supplements for which the proposed target population is children from 1 year onwards and adults. Taking into consideration the composition of the NF and the proposed uses, the consumption of the NF is not nutritionally disadvantageous. There are no concerns regarding genotoxicity of the NF. Based on a 90‐day oral toxicity study conducted with the NF, the Panel considers the NOAEL at the mid‐dose group, i.e. ~ 1000 mg/kg body weight (bw) per day. By applying an uncertainty factor of 200, the resulting intake providing sufficient margin of exposure for humans would be 5 mg/kg bw per day. The available human intervention studies did not report clinically relevant changes in haematological or clinical chemistry parameters following the administration of GH/MGH at supplemental doses of up to 3 g/day for 12 weeks. Overall, the Panel considers that the margin of exposure (~ 200) between the intake of the NF at the proposed uses and use levels and the NOAEL from the 90‐day study is sufficient. The Panel concludes that the NF, glucosyl hesperidin, is safe for the target population at the proposed uses and use levels. [ABSTRACT FROM AUTHOR]

Liquid-liquid phase separation (LLPS) describes many biochemical processes, including hydrogel formation, in the integrity of macromolecular assemblages and existence of membraneless organelles, including ribosome, nucleolus, nuclear speckles, paraspeckles, promyelocytic leukemia (PML) bodies, Cajal bodies (all exert crucial roles in cellular physiology), and evidence are emerging day by day. Also, phase separation is well documented in generation of plasma membrane subdomains and interplay between membranous and membraneless organelles. Intrinsically disordered regions (IDRs) of biopolymers/proteins are the most critical sticking regions that aggravate the formation of such condensates. Remarkably, phase separated condensates are also involved in epigenetic regulation of gene expression, chromatin remodeling, and heterochromatinization. Epigenetic marks on DNA and histones cooperate with RNA-binding proteins through their IDRs to trigger LLPS for facilitating transcription. How phase separation coalesces mutant oncoproteins, orchestrate tumor suppressor genes expression, and facilitated cancer-associated signaling pathways are unravelling. That autophagosome formation and DYRK3-mediated cancer stem cell modification also depend on phase separation is deciphered in part. In view of this, and to linchpin insight into the subcellular membraneless organelle assembly, gene activation and biological reactions catalyzed by enzymes, and the downstream physiological functions, and how all these events are precisely facilitated by LLPS inducing organelle function, epigenetic modulation of gene expression in this scenario, and how it goes awry in cancer progression are summarized and presented in this article. [Display omitted] • LLPS defines macromolecular assemblies, membraneless organelles and biochemical processes. • IDR, LCR, lncRNA and multivalent domain-motif interactions drive LLPS formation. • LLPS regulates chromatin organization and gene expression. • LLPS influence cancer, cancer associated signaling, autophagy and apoptosis. • Machine learning is applicable to predict phase separation and multiple interactions in LLPS. [ABSTRACT FROM AUTHOR]

Copyright of Biology Teaching is the property of East China Normal University and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

The build-up of lactate in solid tumors stands as a crucial and early occurrence in malignancy development, and the concentration of lactate in the tumor microenvironment may be a more sensitive indicator for analyzing primary tumors. In this study, we designed a self-powered lactate sensor for the rapid analysis of tumor samples, utilizing the coupling between the piezoelectric effect and enzymatic reaction. This lactate sensor is fabricated using a ZnO nanowire array modified with lactate oxidase (LOx). The sensing process does not require an external power source or batteries. The device can directly output electric signals containing lactate concentration information when subjected to external forces. The lactate concentration detection upper limit of the sensor is at least 27 mM, with a limit of detection (LOD) of approximately 1.3 mM and a response time of around 10 s. This study innovatively applied self-powered technology to the in situ detection of the tumor microenvironment and used the results to estimate the growth period of the primary tumor. The availability of this application has been confirmed through biological experiments. Furthermore, the sensor data generated by the device offer valuable insights for evaluating the likelihood of remote tumor metastasis. This study may expand the research scope of self-powered technology in the field of medical diagnosis and offer a novel perspective on cancer diagnosis. [ABSTRACT FROM AUTHOR]

Theaflavin-3'-gallate was synthesised using polyphenol oxidase (PPO) from gallocatechin (EGC) and epicatechin gallonic acid (ECG). Using PPO, Box-Behnken design, and single factor test, the optimal reaction conditions were determined: ECG/EGC ratio of 3:7, magnetic stirring speed of 200 rpm, reaction temperature of 37°C, and enzyme concentration of 20 mg/100 mL. Under these conditions, the yield of TF-3'-G was 18.1%. These parameters represent the optimal conversion conditions for theaflavin-3'-monogallate. [ABSTRACT FROM AUTHOR]

Chemical kinetics is a branch of chemistry that investigates the rates of chemical reactions and has applications in cosmology, geology, and physiology. In this study, we develop a mathematical model for chemical reactions based on enzyme dynamics and kinetics, which is a two-step substrate-enzyme reversible reaction, applying chemical kinetics-based modeling of enzyme functions. The non-linear differential equations are transformed into fractional-order systems utilizing the constant proportional Caputo-Fabrizio (CPCF) and constant proportional Atangana-Baleanu-Caputo (CPABC) operators. The system of fractional differential equations is simulated using the Laplace-Adomian decomposition method at different fractional orders through simulations and numerical results. Both qualitative and quantitative analyses such as boundedness, positivity, unique solution, and feasible concentration for the proposed model with different hybrid operators are provided. The stability analysis of the proposed scheme is also verified using Picard's stable condition through the fixed point theorem. [ABSTRACT FROM AUTHOR]

Chemical kinetics is a branch of chemistry that investigates the rates of chemical reactions and has applications in cosmology, geology, and physiology. In this study, we develop a mathematical model for chemical reactions based on enzyme dynamics and kinetics, which is a two-step substrate–enzyme reversible reaction, applying chemical kinetics-based modeling of enzyme functions. The non-linear differential equations are transformed into fractional-order systems utilizing the constant proportional Caputo–Fabrizio (CPCF) and constant proportional Atangana–Baleanu–Caputo (CPABC) operators. The system of fractional differential equations is simulated using the Laplace–Adomian decomposition method at different fractional orders through simulations and numerical results. Both qualitative and quantitative analyses such as boundedness, positivity, unique solution, and feasible concentration for the proposed model with different hybrid operators are provided. The stability analysis of the proposed scheme is also verified using Picard’s stable condition through the fixed point theorem.

Cytochrome P450 monooxygenases (CYPs; P450s) are a superfamily of heme-containing enzymes that are recognized for their vast substrate range and oxidative multifunctionality. CYP107 family members perform hydroxylation and epoxidation processes, producing a variety of biotechnologically useful secondary metabolites. Despite their biotechnological importance, a thorough examination of CYP107 protein structures regarding active site cavity dynamics and key amino acids interacting with bound ligands has yet to be undertaken. To address this research knowledge gap, 44 CYP107 crystal structures were investigated in this study. We demonstrate that the CYP107 active site cavity is very flexible, with ligand binding reducing the volume of the active site in some situations and increasing volume size in other instances. Polar interactions between the substrate and active site residues result in crucial salt bridges and the formation of proton shuttling pathways. Hydrophobic interactions, however, anchor the substrate within the active site. The amino acid residues within the binding pocket influence substrate orientation and anchoring, determining the position of the hydroxylation site and hence direct CYP107's catalytic activity. Additionally, the amino acid dynamics within and around the binding pocket determine CYP107's multifunctionality. This study serves as a reference for understanding the structure–function analysis of CYP107 family members precisely and the structure–function analysis of P450 enzymes in general. Finally, this work will aid in the genetic engineering of CYP107 enzymes to produce novel molecules of biotechnological interest. [ABSTRACT FROM AUTHOR]

Introduction: Cicer arietinum L. is the choice of health food for people with diabetes, hypertension, and hyperlipidemia. As an essential source of high‐nutrition legumes, it is also an important source of dietary isoflavones. Objectives: In order to improve the preparation efficiency of natural plants, a rapid biological activity screening and preparation of xanthine oxidase inhibitors from C. arietinum L. was established. Methods: Xanthine oxidase (XOD) inhibitors were rapidly screened using ultrafiltration liquid chromatography–mass spectrometry (UF‐LC–MS) based on receptor–ligand affinity. The change in XOD activity was evaluated by enzymatic reaction kinetics measurement. The potential bioactive compounds were verified through molecular docking. In addition, the biological activity of ligands screened was separated and purified by complex chromatography. The structures of the compounds were identified by nuclear magnetic resonance spectroscopy. Results: Three active ingredients, namely daidzin, daidzein, calycosin with XOD binding affinities were identified and isolated from the raw plant materials via semi‐preparative high‐performance liquid chromatography (HPLC), 0–60 min, 5–50% B and countercurrent chromatography (CCC) (ethyl acetate/acetic acid/water [5:0.8:10, v/v/v]). Conclusion: This study will help to elucidate the mechanisms of action of natural plants of interest at the molecular level and could also provide more opportunities for the discovery and development of new nutritional value from other natural resources. A novel new techniques for extraction, biological activity screening, and preparation of XOD inhibitors from Cicer arietinum L. have been established. XOD inhibitors were rapidly screened using UF‐LC‐MS. Enzymatic reaction kinetics experiments have been performed to explore its mechanism of action. Meanwhile, the potential anti‐gout effects of the obtained bioactive compounds were verified using molecular docking. Next, three active ingredients, namely, daidzin, daidzein, and calycosin with XOD binding affinities, were identified and isolated by a complex chromatography (semi‐preparative HPLC and CCC). [ABSTRACT FROM AUTHOR]

The viscoelastic behavior of vitreous gel is due to the presence of biopolymers in its structure. Fluid properties of the vitreous is mainly the result of interactions between the characteristics of collagen type II and Hyaluronic Acid networks. Having a better understanding of the structure of each component and their changes during aging and various diseases such as diabetes can lead to better monitoring and treatment options. We study the effects of collagenase type II on 44 samples of porcine vitreous using an in situ rheological experiment in comparison with 18 eyes in a control group injected with Phosphate Buffered Saline Solution. We analyze the behavior of each component over time in both groups. We focus on the changes of viscosity and elasticity of the collagen network within the vitreous. The results of the analysis in this study show that the changes in the fluid properties of the vitreous after collagenase injection is driven by the structural alterations of the collagen network. Creep compliance values of the collagen network are significantly higher in the first group compared to the control group one hour and twenty-four hours after the injection. In contrast, creep compliance of the HA network shows no statistically significant change one hour after the injection in both groups. The results of the reported analysis of individual components in this study support the previous findings on the alterations within the vitreous structure in its entirety.

Amino acids are the most important components of black tea and are associated with the physiological and biochemical changes that occur during tea processing. Herein, we review the enzymatic and non-enzymatic reactions of the main proteinaceous and non-proteinaceous amino acids that occur during the manufacturing process of black tea from the postharvest processing steps of tea leaves to the final product. Enzymatic reactions are responsible for the amino acid variation in the withering, rolling, and fermentation stages. The significant increase in the total amino acid content during the withering stage is mainly derived from proteolysis. Furthermore, the transformation reactions in the rolling and fermentation stages decrease the amino acid levels slightly, whereas the non-enzymatic reactions, such as the Maillard and Strecker reactions, contribute to the significant decrease in the amino acid content of the drying stage. This study provides deep insights into the mechanisms underlying amino acid variation during black tea processing. [ABSTRACT FROM AUTHOR]

A new micro‐kinetic model of the enzyme‐catalyzed synthesis of fructo‐oligosaccharides (FOS) was developed. A commercial enzyme mixture Pectinex® Ultra SP‐L derived from Aspergillus aculeatus was used. A variety of initial enzyme concentrations (1–5 vol%) and sucrose concentrations (400–600 g/L) were experimentally investigated and included in kinetic modeling. Several variations of kinetic mechanisms and corresponding models have been examined. A hybrid genetic algorithm was used to predict the kinetic parameters simultaneously for all experimental data. The best fitting model has been adopted, and with an average error of 13.34%, it describes the experimental data very well. The influence of initial concentrations on the conversion of sucrose and production of FOS is being carefully investigated. It was shown that the initial sucrose concentration significantly affects the highest level of FOS concentration, but the enzyme concentration controls the time at which maximum is reached as well as the rate of FOS decomposition. [ABSTRACT FROM AUTHOR]

An innovative approach based on photothermal contrast to enhance analytical sensitivity of enzymatic phenylalanine detection at a solid surface was demonstrated. Gold nanoparticles in situ produced by the reduced form of nicotinamide adenine dinucleotide, which is generated by an enzymatic reaction, exhibit an excellent photothermal effect upon green-light stimuli. The enzymatic reaction and the nanoparticles process formation were optically investigated. A sensing range of 10–1000 μM with a detection limit of 3.6 μM for phenylalanine detection were demonstrated. The testing with blood samples collected from phenylketonuria patients together with concurrent comparison through tandem mass spectrometry have confirmed the good analytical performances and the robustness and utility of the proposed photothermal-contrast approach for further integration on clinical devices. [ABSTRACT FROM AUTHOR]

A novel fluorescent strategy has been developed by using an enzymatic reaction modulated DNA assembly on graphitic carbon nitride nanosheets (CNNS) for the detection of acetylcholinesterase (AChE) activity and its inhibitors. The two-dimensional and ultrathin-layer CNNS-material was successfully synthesized through a chemical oxidation and ultrasound exfoliation method. Because of its excellent adsorption selectivity to ssDNA over dsDNA and superior quenching ability toward the fluorophore labels, CNNS were employed to construct a sensitive fluorescence sensing platform for the detection of AChE activity and inhibition. The detection was based on enzymatic reaction modulated DNA assembly on CNNS, which involved the specific AChE-catalyzed reaction-mediated DNA/Hg2+ conformational change and subsequent signal transduction and amplification via hybridization chain reaction (HCR). Under the excitation at 485 nm, the fluorescence signal from 500 to 650 nm (λmax = 518 nm) of the developed sensing system was gradually increased with increasing concentration of AChE. The quantitative determination range of AChE is from 0.02 to 1 mU/mL and the detection limit was 0.006 mU/mL. The developed strategy was successfully applied to the assay of AChE in human serum samples, and can also be used to effectively screen AChE inhibitors, showing great promise providing a robust and effective platform for AChE-related diagnosis, drug screening, and therapy. [ABSTRACT FROM AUTHOR]

Enzymatic reactions are crucial to explore the mechanistic function of metabolites and proteins in cellular processes and to understand the etiology of diseases. The increasing number of interconnected metabolic reactions allows the development of in silico deep learning-based methods to discover new enzymatic reaction links between metabolites and proteins to further expand the landscape of existing metabolite–protein interactome. Computational approaches to predict the enzymatic reaction link by metabolite–protein interaction (MPI) prediction are still very limited. In this study, we developed a Variational Graph Autoencoders (VGAE)-based framework to predict MPI in genome-scale heterogeneous enzymatic reaction networks across ten organisms. By incorporating molecular features of metabolites and proteins as well as neighboring information in the MPI networks, our MPI-VGAE predictor achieved the best predictive performance compared to other machine learning methods. Moreover, when applying the MPI-VGAE framework to reconstruct hundreds of metabolic pathways, functional enzymatic reaction networks and a metabolite–metabolite interaction network, our method showed the most robust performance among all scenarios. To the best of our knowledge, this is the first MPI predictor by VGAE for enzymatic reaction link prediction. Furthermore, we implemented the MPI-VGAE framework to reconstruct the disease-specific MPI network based on the disrupted metabolites and proteins in Alzheimer's disease and colorectal cancer, respectively. A substantial number of novel enzymatic reaction links were identified. We further validated and explored the interactions of these enzymatic reactions using molecular docking. These results highlight the potential of the MPI-VGAE framework for the discovery of novel disease-related enzymatic reactions and facilitate the study of the disrupted metabolisms in diseases. [ABSTRACT FROM AUTHOR]

The interest in polymers with high thermal conductivity increased much because of their inherent properties such as low density, low cost, flexibility, and good chemical resistance. However, it is challenging to engineer plastics with good heat transfer characteristics, processability, and required strength. Improving the degree of the chain alignment and forming a continuous thermal conduction network is expected to enhance thermal conductivity. This research aimed to develop polymers with a high thermal conductivity that can be interesting for several applications. Two polymers, namely poly(benzofuran-co-arylacetic acid) and poly(tartronic-co-glycolic acid), with high thermal conductivity containing microscopically ordered structures were prepared by performing enzyme-catalyzed (Novozyme-435) polymerization of the corresponding α-hydroxy acids 4-hydroxymandelic acid and tartronic acid, respectively. A comparison between the polymer's structure and heat transfer obtained by mere thermal polymerization before and enzyme-catalyzed polymerization will now be discussed, revealing a dramatic increase in thermal conductivity in the latter case. The polymer structures were investigated by FTIR spectroscopy, nuclear magnetic resonance (NMR) spectroscopy in liquid- and solid-state (ss-NMR), and powder X-ray diffraction. The thermal conductivity and diffusivity were measured using the transient plane source technique. [ABSTRACT FROM AUTHOR]

Droplet microfluidic chips have emerged as an efficient platform for single-cell analysis due to their high sensitivity, efficiency, and throughput, showing significant potential in pathogen detection. However, current droplet microfluidic chips encounter challenges in large-scale droplet quantification and precise imaging, rendering them unsuitable for the high-throughput pathogen detection required for a large number of samples. To address these issues, this study developed a high-precision fluorescence imaging system utilizing a confocal reflective fluorescence approach, which is an advanced microscopy technique that combines confocal microscopy and reflected fluorescence imaging. It can obtain fluorescence signal and reflected light signal in the sample at the same time, so as to provide richer and more comprehensive image information. The system offers a large field of view (17.8 mm × 17.8 mm) and high resolution (20 μm), enabling the rapid imaging of 30,000 droplets within 10 s, thereby significantly enhancing detection efficiency and automation. Additionally, the enzymatic reaction of Escherichia coli (E. coli) was implemented using the droplet microfluidic chip to validate the effectiveness of the optical imaging system, with results demonstrating the system's capability to accurately capture fluorescence changes during the reaction. Scheme1. Optical system and microfluidic chip structure diagram. [Display omitted] • Large Field Imaging: The system is equipped with a large field fluorescence microscope that rapidly captures large-area droplet images within the microfluidic chip, eliminating the need for image stitching. • High-Throughput Quantification: By integrating image recognition technology, the system analyzes the number of positive and negative E. coli droplets in the chip, enabling fast and precise quantification of E. coli across tens of thousands of droplets. • Rapid Detection: Compared to traditional methods, this system significantly reduces the time required for E. coli detection, enabling near real-time analysis. • Compact Design and Wide Applicability: The system's simplified optical design ensures ease of use and efficient, compact measurement. It is applicable to various biological and medical research fields, providing a highly accurate quantification tool for pathogen detection, genetic analysis, and biomarker research. [ABSTRACT FROM AUTHOR]

Litchi, a tropical and subtropical fruit with substantial commercial value, is prone to browning after harvest. Phytosulfokine (PSK), a plant peptide hormone, plays a significant role in postharvest fruit preservation by affecting various biological processes. However, its specific effects on litchi browning have not been fully elucidated. This study demonstrates that PSK application effectively delays litchi pericarp browning, maintains a higher TSS/TA ratio, and reduces electrical conductivity. PSK treatment inhibits the degradation of anthocyanins, flavonoids, and phenolics, while also decreasing the accumulation of hydrogen peroxide (H₂O₂), malondialdehyde (MDA), and superoxide anions (O₂⁻). Additionally, PSK enhances the activities of superoxide dismutase (SOD), catalase (CAT), and glutathione reductase (GR), while reducing the activities of ascorbate peroxidase (APX), polyphenol oxidase (PPO), and peroxidase (POD) compared to the control. Transcriptome analysis reveals that PSK upregulates genes involved in plant hormone signaling and biosynthesis of anthocyanins and flavonoids, while downregulating genes associated with glutathione metabolism and autophagy. RT-qPCR confirms that PSK treatment decreases the expression levels of APX , LAC7 , PPO , POD1 , POD5 , POD51 , and PODX , while increasing the expression of GPX4 , contributing to the delayed browning. These findings suggest that PSK mitigates litchi browning by modulating oxidative enzymatic reactions and suppressing the expression of browning-related genes. • PSK application delays senescence and reduces pericarp browning in litchi fruits. • PSK boosts SOD, CAT, and GR activities, but reduces APX, PPO, and POD activities. • PSK enhances hormone signaling and anthocyanin biosynthesis genes expression. • PSK downregulates genes linked to glutathione metabolism and autophagy. • PSK delays litchi browning by regulating antioxidant enzymes and browning genes. [ABSTRACT FROM AUTHOR]

Herein, the effect of immobilizing lipase onto layered α-zirconium phosphate ceramic nanosheets (α-ZrP NS) on the enzymatic transesterification of allylic alcohols was investigated in organic solvents. α-ZrP NS modified with quaternary ammonium ions show a excellent dispersion stability in n-hexane. Lipase immobilized on α-ZrP NS enables the improved synthesis of chiral allylic alcohols and esters, with significantly reduced reaction times compared to reactions performed in the absence of α-ZrP NS. The coexistence of α-ZrP NS also results in more efficient steric inversion of the resulting allyl ester, suggesting the utility of α-ZrP NS as a support for lipase-induced stereospecific reactions. The present manuscript discusses on mechanisms of high dispersion stability of α-ZrP NS and their preferable immobilizing effect for the lipase inducing the steric inversion in detail.

In light of the growing interest in products with reduced sugar content, there is a need to consider reducing the natural sugar concentration in juices while preserving the initial concentration of nutritional compounds. This paper reviewed the current state of knowledge related to mixing juices, membrane processes, and enzymatic processes in producing fruit juices with reduced concentrations of sugars. The limitations and challenges of these methods are also reviewed, including the losses of nutritional ingredients in membrane processes and the emergence of side products in enzymatic processes. As the existing methods have limitations, the review also identifies areas that require further improvements and technological innovations. [ABSTRACT FROM AUTHOR]

We created a Microsoft Excel file, Enzyme_Kinetics_Calculator, which includes macro programs that automatically calculates kinetic parameters for typical kinetic equations of enzymatic reactions, accompanied by their standard errors, by minimizing the residual sum of squares thereof. The [>S]-v plot is automatically drawn with the theoretical lines and, similarly, the 1/[5]-1/v plot in the case of linear theoretical lines. Enzyme_Kinetics_Calculator is available as a supplementary file for this paper (see J. Appl. Glycosci. Web site). [ABSTRACT FROM AUTHOR]

Hydrogel fibers are structurally and biologically useful constructs for cell/drug encapsulation as well as fabrication of filament-like tissues. We established cell-laden poly vinyl alcohol (PVA) based hydrogel microfibers through flow focusing microfluidic system and enzymatic crosslinking with desirable cell viability and growth ability. Human adipose stem cell (ADSC)-laden fibers were fabricated by crosslinking phenolic-substituted PVA and gelatin in an aqueous solution containing cells. The crosslinking of phenolic moieties was mediated by horseradish peroxidase reaction and hydrogen peroxide as an electron donor. We have attempted to optimize hydrogel fabrication parameters to obtained microfiber in proper geometries and dimensions. The producing PVA/Gelatin microfibers could be rolled and produced bundled filament-like structure. The physical characteristics of produced microfibers were evaluated. The hybrid PVA/Gelatin microfiber is shown 0.18 swelling ratio which was lower than PVA sample. While, the tensile strength and elongation of PVA/Gelatin derivative significantly upregulated and demonstrated 413 kPa tensile strength and 4.1 times elongation. The encapsulated ADSCs proliferated and grew within hydrogel microfibers and formed filament-like cellular constructs in single and bundled constructs. These results demonstrate the feasibility of PVA-based hydrogel fibers obtained through flow focusing microfluidic system and HRP-mediated crosslinking reaction for the engineering of filament-like tissues. [Display omitted] • Poly vinyl alcohol (PVA) based hydrogel microfibers fabricated through flow focusing microfluidic system. • PVA-based microfibers possessed proper geometries and dimensions in optimum operational condition. • Mechanical properties of PVA/Gelatin significantly improved and demonstrated higher tensile strength and elongation. • Encapsulated cells grew and proliferated within PVA/Gelatin hydrogel microfibers. [ABSTRACT FROM AUTHOR]

Visualizing the distribution of enzymes is vital for understanding physiological phenomena. Enzyme histochemistry is a technique used to investigate the localization of enzyme activity. However, the target is restricted to enzymes with easy-to-design artificial substrates that can develop color through reactions. Mass spectrometry imaging (MSI)-based enzyme histochemistry has been developed as a novel method to visualize enzyme localization. It can be applied to enzyme histochemistry as it detects products from the supplied substrate using enzymes present on the tissue sections. However, enzyme histochemistry using MSI has not been applied to plant tissue samples yet. Glutamate decarboxylase (GAD, EC: 4.1.1.15) is an enzyme that catalyzes the decarboxylation reaction of l -glutamic acid to produce γ-aminobutyric acid (GABA). GABA biosynthesis is important both in the field of food chemistry and plant physiology. This study focused on GAD during the legume germination process and successfully visualized GAD activity in legume seeds using MSI for the first time. Furthermore, the localization of GAD activity in the embryonic axis of germinated soybean seeds and alfalfa seeds could be visualized. • Mass spectrometry imaging was used to visualize enzymatic activity on plant samples for the first time. • Enzyme localization was visualized using MSI in soybean and alfalfa seeds. • Highest enzyme activity detected at 60 °C at a pH of 5.8. • GAD activity was highest in germinated seeds compared to non-germinated seeds. • Enzymes had a constant reaction rate when sufficient substrate was available. [ABSTRACT FROM AUTHOR]

Trichosanthes kirilowii Maxim taxonomically belongs to the Cucurbitaceae family and Trichosanthes genus. Its whole fruit, fruit peel, seed and root are widely used in traditional Chinese medicines. A ribosome-inactivating protein with RNA N-glycosidase activity called Trichosanthrip was isolated and purified from the seeds of T. kirilowii in our recent previous research. To further explore the biological functions of Trichosanthrip, the cDNA of T. kirilowii alpha-amylase inhibitor (TkAAI) was cloned through rapid-amplification of cDNA ends and its sequence was analyzed. Also, the heterologous protein was expressed in Escherichia coli and its alpha-amylase activity was further measured under optimized conditions. The full-length cDNA of TkAAI was 613 bp. The speculated open reading frame sequence encoded 141 amino acids with a molecular weight of 16.14 kDa. Phylogenetic analysis demonstrated that the Alpha-Amylase Inhibitors Seed Storage domain sequence of TkAAI revealed significant evolutionary homology with the 2S albumin derived from the other plants in the Cucurbitaceae group. In addition, TkAAI was assembled into pET28a with eGFP to generate a prokaryotic expression vector and was induced to express in E. coli. The TkAAI-eGFP infusion protein was proven to exhibit alpha-amylase inhibitory activity against porcine pancreatic amylase in a suitable reaction system. Analysis of gene expression patterns proved that the relative expression level of TkAAI in seeds is highest. The results presented here forecasted that the TkAAI might play a crucial role during the development of T. kirilowii seeds and provided fundamental insights into the possibility of T. kirilowii derived medicine to treat diabetes related diseases. [ABSTRACT FROM AUTHOR]

The longan pericarp turns brown dramatically after harvesting, but the mechanism is not well understood. In this work, two peroxidases were purified from longan pericarp and found to be identical to the class III peroxidases PRX53-2 and PRX53-3. In vitro , PRX53-2/3 catalyzed the browning of several pericarp abundant proanthocyanidin and lignin monomers, such as (−)-epicatechin (EC), (+)-catechin (CT) and coniferyl alcohol (ConA). PRX53–2 was upregulated and highly-expressed, while PRX53-3 was expressed at low levels after harvesting; thus, PRX53-2 was considered a browning-related gene. The reaction with both proanthocyanidin and lignin presented a greater degree of brown coloration compared to the single substrate reactions. Several procyanidins isomers, EC-ConA and CT-ConA were detected in the double-substrate reaction. These results not only demonstrate that the effects of PRX53-2 on proanthocyanidin and lignin polymerization may be crucial for longan pericarp browning, but also help in developing new strategies or preservatives to delay pericarp browning. • PRX53-2/3 was purified from longan pericarp and showed high activity to guaiacol. • PRX53-2 is highly expressed during longan pericarp browning in postharvest storage. • PRX53-2 catalyzes proanthocyanidin and lignin monomers polymerization in vitro. • The polymerization of (epi)catechin is enhanced when ConA is present. • A new proposed mechanism helps in developing strategies to delay pericarp browning. [ABSTRACT FROM AUTHOR]

Inspired by the natural world's fascinating capacity for color change, researchers have developed various responsive hydrogels that shift colors. Among these, hydrogels that react to pH changes are particularly notable for their unique properties. The urea-urease autocatalytic reaction, a key pH clock reaction, is especially effective in temporarily adjusting these hydrogels' properties and enabling color changes in response to stimuli. In this study, we synergize the urea-urease clock reaction with the pH-sensitive color transformation of neutral red (NR), thereby establishing an NR-urea-urease clock reaction system. This system exhibits a dynamic color cycle from yellow to red and back to yellow. By incorporating this color transition mechanism into polymer hydrogels, we successfully initiate a pH change within the system. This innovation paves the way for a smart material capable of displaying temporary information through enzymatic reactions. The temporal variation in information display is finely tuned by adjusting the urea and urease concentrations. Such reversible color-changing hydrogels hold promising prospects for applications in secure information transmission and dynamic camouflage, marking a significant step forward in systems chemistry and materials science. [Display omitted] • This study develops reversible, color-changing hydrogels for dynamic displays. • Smart hydrogels enable secure information transmission and adaptive camouflage. • Optimized hydrogel formulations enhance water retention and mechanical strength. • A novel method for on-demand encoding and erasing of information is introduced. [ABSTRACT FROM AUTHOR]

Methicillin-resistant Staphylococcus aureus (MRSA) is a multidrug-resistant bacterium that causes a wide range of illnesses, necessitating the development of new technologies for its detection. Herein, we propose a graphene oxide (GO)-based sensing platform for the detection of mecA gene in MRSA using flap endonuclease 1 (FEN1)-assisted target recycling and Klenow fragment (KF)-triggered signal amplification. Without the target, all the DNA probes were adsorbed onto GO, resulting in fluorescence quenching of the dye. Upon the addition of the target, a triple complex was formed that triggered FEN1-assisted target recycling and initiated two polymerization reactions with the assistance of KF polymerase, generating numerous dsDNA that were repelled by GO. These dsDNAs triggered fluorescence enhancement when SYBR Green I was added. Therefore, the target DNA was quantified by measuring the fluorescence at excitation and emission wavelengths of 480/526 nm. This mecA gene assay showed a good linear range from 1 to 50 nM with a lower limit of detection of 0.26 nM, and displayed good applicability to the analysis of real samples. Thus, a new method for monitoring MRSA has been developed that has great potential for early clinical diagnosis and treatment. • A novel GO-based bioassay was constructed for efficient assay of mecA gene in MRSA. • Integration of FEN1 and KF in one system that trigger signal amplification to improve the bioassay performance. • The bioassay can achieve low detection limits of 0.26 nM within 5 min. • The bioassay was successfully applied to determination of MRSA in real samples. [ABSTRACT FROM AUTHOR]

The surface droplet evaporation-based sensing platform for the screening of a lipase inhibitor orlistat based on measuring the area of aqueous droplet from the lipase-mediated enzymatic reaction. [Display omitted] • A surfactant-mediated surface droplet evaporation-based biosensor is developed. • It shows excellent performance for screening lipase inhibitors, with an IC 50 value of 0.59 ± 0.09 μM for orlistat. • The sodium oleate produced by lipase digestion of triacylglycerols reduces surface tension and increases the dried area on the surface. • Olistat inhibits lipase activity and reduces the dried area on the surface. • It also works well with a portable microscope and a smartphone interface for data analysis. The development of new types of sensing platforms with excellent performance is always appealing. In this report, we demonstrate a low-cost and user-friendly sensing platform based on drying of the aqueous droplet on the surface. The principle relies on the transduction of the surfactant concentration into the area of the dried droplet on the surface. The screening of a lipase inhibitor is demonstrated as an example. When triacylglycerols (GT) and lipase are mixed, the aqueous droplet produces a large dried area on the surface. This phenomenon is attributed to the production of sodium oleate due to the enzymatic digestion of GT by lipase. It results in reduction of the surface tension and causes wetting and spreading of the droplet, leaving a large dried area on the surface. However, in the presence of orlistat, a lipase inhibitor, a small dried area is obtained due to the inhibition of lipase activity by orlistat. Using this method, the lipase inhibitor can be readily and effectively screened. Compared to previous studies, this method is simple, rapid, low-cost and avoids the use of labelled molecules and complex instrumentation, with an IC 50 value of 0.59 ± 0.09 μM for orlistat. In addition, it also works well with a portable microscope and a smartphone interface for data analysis. Overall, this study provides a new method for rapid and cost-effective screening of lipase inhibitors. [ABSTRACT FROM AUTHOR]

A liquid crystal (LC)-based assay was developed to detect chlorothalonil (CHL). The detection principle is based on (i) the electrostatic interaction between the positively charged protein protamine (PRO) with the negatively charged phospholipid dioleoyl-sn-glycero-3-phospho-rac-(1-glycerol) sodium salt (DOPG) and (ii) the CHL-mediated inhibition of papain (PAP) activity. The aqueous/LC interface was decorated with a monolayer of DOPG and PRO that self-assembled via electrostatic interactions. PAP can hydrolyze PRO, resulting in the realignment of an LC by DOPG, inducing a shift in the LC response from bright to dark. The addition of CHL can inhibit the activity of PAP, leading to the attraction of PRO to DOPG and the consequent disruption of the LC orientation. The orientation change of the LC in the presence or absence of CHL can be observed from the changes in its optical appearance using a polarized light microscope. Under optimal conditions, the developed assay achieved a detection limit of 0.196 pg mL−1 within a range of determination of 0.65–200 pg mL−1. The selectivity of the assay was verified in the presence of carbendazim and imidacloprid. The practical application of the proposed assay was demonstrated by its use to determine the levels of CHL in food extracts and environmental samples, which yielded recoveries and relative standard deviations (RSD) in the ranges of 87.39–99.663% and 1.03–6.32%, respectively. [ABSTRACT FROM AUTHOR]

Enzyme histochemistry via mass spectrometry imaging (MSI) has garnered attention as a straightforward approach for visualizing enzymatic reactions. While several studies in the medical and physiological fields have shown its promising application potential, its applicability to agricultural or food studies has not yet been demonstrated. Rice koji , known as an enzyme source for various fermented products, is a suitable model for demonstrating the applicability of this method to food-related materials. In this study, the enzymatic reaction of dipeptidyl peptidase B (DppB) in rice koji was visualized using MSI for the first time. The method was optimized and applied to investigate the effects of rice variety, polishing ratio, and cultivation time on the location of the DppB reaction. The DppB enzymatic reaction was found to occur in different locations in each of the two rice varieties, Yamadanishiki and Hakutsurunishiki. The polishing ratio also affected the distribution of the DppB enzymatic reactions. Furthermore, a time-course investigation of rice koji cultivation revealed that while the location of the reaction was largely associated with mycelial penetration, the structure and features of the rice grain may also affect the location of the enzymatic reaction. In summary, these results demonstrate the applicability of enzyme histochemistry by MSI to food-related materials. • Applicability of mass spectrometry imaging (MSI) to food analysis was studied. • The enzymatic activity of dipeptidyl peptidase B in rice koji was analyzed via MSI. • The method was optimized. • The effects of rice variety, polishing ratio, and cultivation time were studied. • MSI was demonstrated to be applicable for food-material analysis. [ABSTRACT FROM AUTHOR]

Abstract In this paper, the authors present a strategy based on fixed point iterative methods to solve a nonlinear dynamical problem in a form of Green’s function with boundary value problems. First, the authors construct the sequence named Green’s normal-S iteration to show that the sequence converges strongly to a fixed point, this sequence was constructed based on the kinetics of the amperometric enzyme problem. Finally, the authors show numerical examples to analyze the solution of that problem.

SNAP-tag® is a powerful technology for the labelling of protein/enzymes by using benzyl-guanine (BG) derivatives as substrates. Although commercially available or ad hoc produced, their synthesis and purification are necessary, increasing time and costs. To address this limitation, here we suggest a revision of this methodology, by performing a chemo-enzymatic approach, by using a BG-substrate containing an azide group appropriately distanced by a spacer from the benzyl ring. The SNAP-tag® and its relative thermostable version (SsOGT-H5) proved to be very active on this substrate. The stability of these tags upon enzymatic reaction makes possible the exposition to the solvent of the azide-moiety linked to the catalytic cysteine, compatible for the subsequent conjugation with DBCO-derivatives by azide-alkyne Huisgen cycloaddition. Our studies propose a strengthening and an improvement in terms of biotechnological applications for this self-labelling protein-tag.

Herein, the effect of immobilizing lipase onto layered α-zirconium phosphate ceramic nanosheets (α-ZrP NS) on the enzymatic transesterification of allylic alcohols was investigated in organic solvents. α-ZrP NS modified with quaternary ammonium ions show a excellent dispersion stability in n-hexane. Lipase immobilized on α-ZrP NS enables the improved synthesis of chiral allylic alcohols and esters, with significantly reduced reaction times compared to reactions performed in the absence of α-ZrP NS. The coexistence of α-ZrP NS also results in more efficient steric inversion of the resulting allyl ester, suggesting the utility of α-ZrP NS as a support for lipase-induced stereospecific reactions. The present manuscript discusses on mechanisms of high dispersion stability of α-ZrP NS and their preferable immobilizing effect for the lipase inducing the steric inversion in detail. [ABSTRACT FROM AUTHOR]