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A dinuclear copper (II) complex, Cu2 (RCOO)4(L)2 (1) while [RCOO = benzoate, L = 2-amino-3-chloro-5-trifluoro methyl pyridine], with square pyramidal geometry with four carboxylate bridged groups has been reported. This dinuclear copper (II) unit (with the most robust, frequently occurring paddle wheel structure) was characterized by single crystal X-ray diffraction, IR, UV, CV, TGA, and EPR studies. It exhibits catechol oxidase activity in methanol medium and the catecholase activity was monitored by the UV–Vis spectroscopy. The kinetic parameters have been determined by using Michaelis–Menten equation which shows that the complex is an efficient catalyst with very high turnover number. Preliminary mechanistic investigation of the catalytic behavior was established with the help of ESI–MS spectra. [ABSTRACT FROM AUTHOR]

Modified atmosphere storage containers with gas-permeable membranes are effective in reducing post-harvest losses, mainly at low temperatures but become ineffective in controlling O 2 and CO 2 at temperatures above 3 °C. This study aimed to develop a method to control O 2 in a storage box. This method uses a diffusion tube, that blocks air from entering the box but allows for air exchange when an air blower is activated. The blower ON frequency (BOF) was modelled as a function of temperature, considering the type and amount of produce, blower properties, tube dimension, and the O 2 setpoint. The effects of temperature and gas composition on respiration rate kinetics were analysed with Michaelis-Menten and Arrhenius equations. The model was used to program the microcontroller to control the blower. Validation was performed using a 190 L box containing 25 kg of broccoli. The developed model maintained the desired O 2 concentration under fluctuating temperatures. The BOF values ranged from 25.6 to 549.2 s h−1, for temperature fluctuations between 4 and 23 °C. The time required to reach the 3% O 2 in the box differed depending on the blower's state. With the blower turned ON from the beginning, it took 24.9 h. However, when the blower was initially turned OFF, it took 11.1 h. Despite temperature changes, the system maintained O 2 at 3.8 ± 0.29% and CO 2 at 14.4 ± 0.66%. The system is promising for commercial use and best suited for CO 2 tolerant produce because it requires a separate mathematical model and control mechanism. • Simple method to control gas concentrations under fluctuating temperature. • Real-time O 2 control in storage based on temperature sensor and predictive model. • Mathematical model guides real-time air-blower activation for maintaining O 2. • Suitable for CO 2 -tolerant produce or when combined with CO 2 removal tools. [ABSTRACT FROM AUTHOR]

The chemical etching of germanium in Br2 environment at elevated temperatures is described by the Michaelis–Menten equation. The validity limit of Michaelis–Menten kinetics is subjected to the detailed analysis. The steady-state etching rate requires synergy of two different process parameters. High purity gas should be directed to the substrate on which intermediate reaction product does not accumulate. Theoretical calculations indicate that maximum etching rate is maintained when 99.89% of the germanium surface is covered by the reaction product, and 99.9999967% of the incident Br2 molecules are reflected from the substrate surface. Under these conditions, single GeBr2 molecule is formed after 30 million collisions of Br2 molecules with the germanium surface. [ABSTRACT FROM AUTHOR]

Background: Standard pediatric growth curves cannot be used to impute missing height or weight measurements in individual children. The Michaelis–Menten equation, used for characterizing substrate-enzyme saturation curves, has been shown to model growth in many organisms including nonhuman vertebrates. We investigated whether this equation could be used to interpolate missing growth data in children in the first three years of life and compared this interpolation to several common interpolation methods and pediatric growth models. Methods: We developed a modified Michaelis–Menten equation and compared expected to actual growth, first in a local birth cohort (N = 97) then in a large, outpatient, pediatric sample (N = 14,695). Results: The modified Michaelis–Menten equation showed excellent fit for both infant weight (median RMSE: boys: 0.22 kg [IQR:0.19; 90% < 0.43]; girls: 0.20 kg [IQR:0.17; 90% < 0.39]) and height (median RMSE: boys: 0.93 cm [IQR:0.53; 90% < 1.0]; girls: 0.91 cm [IQR:0.50;90% < 1.0]). Growth data were modeled accurately with as few as four values from routine well-baby visits in year 1 and seven values in years 1–3; birth weight or length was essential for best fit. Interpolation with this equation had comparable (for weight) or lower (for height) mean RMSE compared to the best performing alternative models. Conclusions: A modified Michaelis–Menten equation accurately describes growth in healthy babies aged 0–36 months, allowing interpolation of missing weight and height values in individual longitudinal measurement series. The growth pattern in healthy babies in resource-rich environments mirrors an enzymatic saturation curve. [ABSTRACT FROM AUTHOR]

The kinetically-derived maximal dose (KMD) is defined as the maximal external dose at which kinetics are unchanged relative to lower doses, e.g., doses at which kinetic processes are not saturated. Toxicity produced at doses above the KMD can be qualitatively different from toxicity produced at lower doses. Here, we test the hypothesis that neoplastic lesions reported in the National Toxicology Program's (NTP) rodent cancer bioassay with ethylbenzene are a high-dose phenomenon secondary to saturation of elimination kinetics. To test this, we applied Bayesian modeling on kinetic data for ethylbenzene from rats and humans to estimate the Vmax and Km for the Michaelis–Menten equation that governs the elimination kinetics. Analysis of the Michaelis–Menten elimination curve generated from those Vmax and Km values indicated KMD ranges for venous ethylbenzene of 8–17 mg/L in rats and 10–18 mg/L in humans. Those venous concentrations are produced by inhalation concentrations of around 200 ppm ethylbenzene, which is well above typical human exposures. These KMD estimates support the hypothesis that neoplastic lesions seen in the NTP rodent bioassay occur secondary to saturation of ethylbenzene elimination pathways and are not relevant for human risk assessment. Thus, ethylbenzene does not pose a credible cancer risk to humans under foreseeable exposure conditions. Cancer risk assessments focused on protecting human health should avoid endpoint data from rodents exposed to ethylbenzene above the KMD range and future toxicological testing should focus on doses below the KMD range. [ABSTRACT FROM AUTHOR]

Desiccation is a severe survival problem for organisms. We have been studying the desiccation tolerance mechanisms in the true slime mold Physarum polycephalum. We measured the trehalose content of P. polycephalum vegetative cells (plasmodia) and drought cells (sclerotia). Surprisingly, we found that the content in sclerotia was about 473-fold greater than in the plasmodia. We then examined trehalose metabolism-related genes via RNAseq, and consequently found that trehalose 6-phosphate phosphorylase (T6pp) expression levels increased following desiccation. Next, we cloned and expressed the genes for T6pp, trehalose 6-phosphate synthase/phosphatase (Tps/Tpp), maltooligosyltrehalose trehalohydrolase (TreZ), and maltooligosyltrehalose synthase (TreY) in E. coli. Incidentally, TreY and TreZ clones have been reported in several prokaryotes, but not in eukaryotes. This report in P. polycephalum is the first evidence of their presence in a eukaryote species. Recombinant T6pp, TreY, and TreZ were purified and confirmed to be active. Our results showed that these enzymes catalyze reactions related to trehalose production, and their reaction kinetics follow the Michaelis-Menten equation. The t6pp mRNA levels of the sclerotia were about 15-fold higher than in the plasmodia. In contrast, the expression levels of TreZ and TreY showed no significant change between the sclerotia and plasmodia. Thus, T6pp is probably related to desiccation tolerance, whereas the contribution of TreY and TreZ is insufficient to account for the considerable accumulation of trehalose in sclerotia. • Study for the desiccation tolerance mechanisms in a true slime mold Physarum polycephalum. • A large amount of trehalose was accumulated in the drought cells (sclerotia). • The genes (t6pp , tps/tpp , treZ , treY) related to trehalose metabolism were cloned. • T6pp is probably related to desiccation tolerance. • This is the first report of treY and treZ genes from eukaryotic species. [ABSTRACT FROM AUTHOR]

The microbial community composition is crucial for metal extraction from solid waste by bioleaching. However, bioreactor output of bioleachant with a diverse microbial population is a great challenge due to the different growth conditions of each species. This study reports an immobilized bioreactor for the cultivation of mesophilic consortia. The optimal conditions for iron oxidation were pH 1.5, dilution of 0.382 h–1, the temperature of 30°C, and 12 g/L ferrous iron. The coexistence relationship of Acidithiobacillus ferrooxidans, Acidithiobacillus ferrivorans, and Acidiphilium spp. was relatively stable in a wider range of conditions. Further kinetics investigation showed that ferrous iron oxidation fitted well with the substrate free inhibition model of Michaelis−Menten equation below 12 g/L ferrous iron, indicating that the immobilized bioreactor culturing mesophilic consortia slowed down the inhibition process caused by the substrate and end-product. These key features would enable bioreactor design more efficient during industrial applications of biolea-ching. [ABSTRACT FROM AUTHOR]

It is crucial for hemostasis that platelets are rapidly recruited to the site of vascular injury by the adhesive ligand von Willebrand factor (VWF) multimers. The metalloproteinase ADAMTS13 regulates this hemostatic activity by proteolytically reducing the size of VWF and its proteolytic kinetics has been investigated by biochemical and single-molecule biophysical methods. However, how ADAMTS13 cleaves VWF in flowing blood remains poorly defined. To investigate the force-induced VWF cleavage, VWF A1A2A3 tridomains were immobilized and subjected to hydrodynamic forces in the presence of ADAMTS13. We demonstrated that the cleavage of VWF A1A2A3 by ADAMTS13 exhibited biphasic kinetics governed by shear stress, but not shear rate. By fitting data to the single-molecule Michaelis-Menten equation, the proteolytic constant k cat of ADAMTS13 had two distinct states. The mean proteolytic constant of the fast state (k cat-fast) was 0.005 ± 0.001 s−1, which is >10-fold faster than the slow state (k cat-slow = 0.0005 ± 0.0001 s−1). Furthermore, proteolytic constants of both states were regulated by shear stress in a biphasic manner, independent of the solution viscosity, indicating that the proteolytic activity of ADAMTS13 was regulated by hydrodynamic force. The findings provide new insights into the mechanism underlying ADAMTS13 cleaving VWF under flowing blood. • The cleavage of VWF A1A2A3 by ADAMTS13 exhibits a biphasic manner under flow. • ADAMTS13 cleaving VWF A1A2A3 showed fast and slow proteolytic states. • Both states were biphasically regulated by shear stress, implying an optimal configuration of unfolded A2 for cleavage. [ABSTRACT FROM AUTHOR]

Many diseases in the human body are related to the level of L-cysteine. Therefore, it is crucial to establish an efficient, simple and sensitive platform for L-cysteine detection. In this work, we synthesized platinum palladium bimetallic nanoparticles (Van-Ptm/Pdn NPs) using vancomycin hydrochloride (Van) as a stabilizer, which exhibited high oxidase-like catalytic activity. In addition, the catalytic kinetics of the Van-Pt1/Pd1 NPs followed the typical Michaelis–Menten equation, exhibiting a strong affinity for 3,3′,5,5′-tetramethylbenzidine substrates. More importantly, we developed a simple and effective strategy for the sensitive colorimetric detection of L-cysteine using biocompatible Van-Pt1/Pd1 NPs. The detection limit was low, at 0.07 μM, which was lower than the values for many previously reported enzyme-like detection systems. The colorimetric method of the L-cysteine assay had good selectivity. The established method for the detection of L-cysteine showed promise for biomedical analysis. [ABSTRACT FROM AUTHOR]

Rhyzopertha dominica is a pest that uses trypsin-like serine protease enzymes to hydrolyse the proteins in the cereal grains on which it feeds. The present study reveals for the first time that that there are both glycosylated and non-glycosylated serine proteases. The progeny of R. dominica reared on the grain three varieties of wheat were used to fractionate their trypsinlike serine proteases using Concanavalin A affinity chromatography. The albumin fractions from the wheat cultivars used in this study were subjected to size exclusion chromatography to fractionate the albumin inhibitors that are highly specific for the serine protease activity of R. dominica. Kinetic and thermodynamic assays were used to differentiate both types of enzymes. In general, the catalytic efficiency values Vmax/Km for glycosylated proteases were higher, indicating that glycosylation increases the affinity for the substrate. Inhibition assays using wheat albumins revealed that the glycosylated enzymes had higher Ki values, indicating a low affinity for the inhibitors than the non-glycosylated enzymes. Thermodynamic analysis indicates that glycosylation increases the activation energy Ea improving the serine proteases' catalysis. Thus it is likely that R. dominica uses glycosylated proteases in order to optimize the hydrolysis of cereal proteins and nullify the action of wheat grain protease inhibitors and increase its chances of survival. [ABSTRACT FROM AUTHOR]

The hyperbolic dependence of catalytic rate on substrate concentration is a classical result in enzyme kinetics, quantified by the celebrated Michaelis–Menten equation. The ubiquity of this relation in diverse chemical and biological contexts has recently been rationalized by a graph-theoretic analysis of deterministic reaction networks. Experiments, however, have revealed that "molecular noise"—intrinsic stochasticity at the molecular scale—leads to significant deviations from classical results and to unexpected effects like "molecular memory," i.e., the breakdown of statistical independence between turnover events. Here, we show, through a new method of analysis, that memory and non-hyperbolicity have a common source in an initial, and observably long, transient peculiar to stochastic reaction networks of multiple enzymes. Networks of single enzymes do not admit such transients. The transient yields, asymptotically, to a steady-state in which memory vanishes and hyperbolicity is recovered. We propose new statistical measures, defined in terms of turnover times, to distinguish between the transient and steady-states and apply these to experimental data from a landmark experiment that first observed molecular memory in a single enzyme with multiple binding sites. Our study shows that catalysis at the molecular level with more than one enzyme always contains a non-classical regime and provides insight on how the classical limit is attained. [ABSTRACT FROM AUTHOR]

This study reports the preparation of silica-coated and nano-fructosome encapsulated Candida antarctica lipase B particles (CalB@NF@SiO2) and a demonstration of their enzymatic hydrolysis and acylation. CalB@NF@SiO2 particles were prepared as a function of TEOS concentration (3–100 mM). Their mean particle size was 185 nm by TEM. Enzymatic hydrolysis was performed to compare catalytic efficiencies of CalB@NF and CalB@NF@SiO2. The catalytic constants (Km, Vmax, and Kcat) of CalB@NF and CalB@NF@SiO2 were calculated using the Michaelis–Menten equation and Lineweaver–Burk plot. Optimal stability of CalB@NF@SiO2 was found at pH 8 and a temperature of 35 °C. Moreover, CalB@NF@SiO2 particles were reused for seven cycles to evaluate their reusability. In addition, enzymatic synthesis of benzyl benzoate was demonstrated via an acylation reaction with benzoic anhydride. The efficiency of CalB@NF@SiO2 for converting benzoic anhydride to benzyl benzoate by the acylation reaction was 97%, indicating that benzoic anhydride was almost completely converted to benzyl benzoate. Consequently, CalB@NF@SiO2 particles are better than CalB@NF particles for enzymatic synthesis. In addition, they are reusable with high stability at optimal pH and temperature. [ABSTRACT FROM AUTHOR]

Hydrogen peroxide (H 2 O 2) is an important oxidant and one common active oxygen species in organisms. It is critically important to realize fast and accurate detection of H 2 O 2. In this work, Cu–SnO 2 is successfully evaluated as a dual mode-sensing material for H 2 O 2 detection. The study proves that the peroxidase-like activity increases steadily with the oxygen vacancy concentration on the material surface. The material Cu–SnO 2 -1.5, which shows the highest amount of oxygen vacancy, successfully oxidizes colorless substrate (TMB) to blue (ox TMB) in the presence of H 2 O 2. The peroxidase-like behaviors of this material are consistent with the Michaelis-Menten equation. An electrochemical sensor based on this material exhibits a wide working range (0.2 μM–0.745 mM and 0.745–5.345 mM), a low detection limit of 67 nM, and promising sensitivity of 158.16 μA·mM−1cm−2 for H 2 O 2 in 0.1 M phosphate buffered (PB) solution. In addition, the H 2 O 2 released from human liver cancer (HepG2) cells is successfully detected by the sensor, proving its potential application in situ H 2 O 2 detection. [ABSTRACT FROM AUTHOR]

Food-derived antihypertensive peptides have attracted increasing attention in functional foods for health promotion, due to their high biological activity, low toxicity and easy metabolism in the human body. Angiotensin converting enzyme (ACE) is a key enzyme that causes the increase in blood pressure in mammals. However, few reviews have summarized the current understanding of ACE inhibitory peptides and their knowledge gaps. This paper focuses on the food origins and production methods of ACE inhibitory peptides. Compared with conventional methods, the advanced technologies and emerging bioinformatics approaches have recently been applied for efficient and targeted release of ACE inhibitory peptides from food proteins. Furthermore, the transport and underlying mechanisms of ACE inhibitory peptides are emphatically described. Molecular modeling and the Michaelis-Menten equation can provide information on how ACE inhibitors function. Finally, we discuss the structure-activity relationships and other bio-functional properties of ACE inhibitory peptides. Molecular weight, hydrophobic amino acid residues, charge, amino acid composition and sequence (especially at the C-terminal and N-terminal) have a significant influence on ACE inhibitory activity. Some studies are required to increase productivity, improve bioavailability of peptides, evaluate their bio-accessibility and efficiency on reducing blood pressure to provide a reference for the development and application of health products and auxiliary treatment drugs. [ABSTRACT FROM AUTHOR]

Exploring highly active peroxidase mimics at physiological pH is important for the construction of efficient and convenient colorimetric sensing platforms for detecting small biomolecules. In this work, prepared zinc pyrovanadate (Zn3V2O7(OH)2·2H2O) nanorods exhibit excellent peroxidase-like activity, which is verified by the fast oxidation of colorless 3,3′,5,5′-tetramethylbenzidine (TMB) into a blue product (oxTMB) by H2O2 at physiological pH (pH = 7) in 2 min. In addition, the catalytic behaviors of Zn3V2O7(OH)2·2H2O as a peroxidase-like nanozyme conform to the Michaelis–Menten equation. Scavenger experiments prove that the catalytic activity of Zn3V2O7(OH)2·2H2O is ascribed to ˙O2− radicals generated in the process of catalysis. Based on the peroxidase-like activity of the Zn3V2O7(OH)2·2H2O nanozyme, a fast and convenient colorimetric sensor has been constructed to detect H2O2 and epinephrine (EP) under physiological pH. The detection limit of EP is as low as 0.26 μM. In addition, the feasibility of the proposed sensor has been validated to detect H2O2 in milk and EP in serum. [ABSTRACT FROM AUTHOR]

In this study, a heterogeneous multiscale method based mass transport model is developed for delivering drug into biological tissues. In the model, fluid flow and drug metabolism are modelled at the continuum scale, whereas the diffusion phenomenon is incorporated through macro-micro coupling. Darcy's law is used for fluid flow calculation, and the drug metabolism is determined using the Michaelis-Menten equation. Using the model, penetration and distribution of a drug in a tissue are investigated. Mainly, the effects of fluid flow and drug metabolism together with the particle size effects on drug delivery are explored. It is observed that the particles of sizes 10 and 100 nm can penetrate the tissue with advection; however, without advection, such particles accumulate around the capillary, the source of the drug to the interstitium. In fluid flow regions, drug metabolism may not affect penetration and distribution of drug; the results are similar to those without drug metabolism. However, metabolism may significantly reduce drug penetration in the absence of fluid flow. It is observed from the sensitivity analysis that the model is most sensitive to the parameters that model the transport processes of extracellular space while least sensitive to those that incorporate the intracellular processes. [ABSTRACT FROM AUTHOR]

Analyzing the molecular and physiological processes that govern the uptake and transport of nitrogen (N) in plants is central to efforts to fully understand the optimization of plant N use and the changes in the N-use efficiency in relation to changes in atmospheric N deposition changes. Here, a field experiment was conducted using the ectomycorrhizal fungi (EMF), Pisolithus tinctorius (Pt) and Suillus grevillei (Sg). The effects of N deposition were investigated using concentrations of 0 kg·N·hm−2a−1 (N0), a normal N deposition of 30 kg·N·hm−2a−1 (N30), a moderate N deposition of 60 kg·N·hm−2a−1 (N60), and a severe N deposition of 90 kg·N·hm−2a−1 (N90), with the goal of examining how these factors impacted root activity, root absorbing area, NH4+ and NO3− uptake kinetics, and the expression of ammonium and nitrate transporter genes in Pinus massoniana seedlings under different levels of N deposition. These data revealed that EMF inoculation led to increased root dry weight, activity, and absorbing area. The NH4+ and NO3− uptake kinetics in seedlings conformed to the Michaelis–Menten equation, and uptake rates declined with increasing levels of N addition, with NH4+ uptake rates remaining higher than NO3− uptake rates for all tested concentrations. EMF inoculation was associated with higher Vmax values than were observed for non-mycorrhizal plants. Nitrogen addition resulted in the upregulation of genes in the AMT1 family and the downregulation of genes in the NRT family. EMF inoculation under the N60 and N90 treatment conditions resulted in the increased expression of each of both these gene families. NH4+ and NO3− uptake kinetics were also positively correlated with associated transporter gene expression in P. massoniana roots. Together, these data offer a theoretical foundation for EMF inoculation under conditions of increased N deposition associated with climate change in an effort to improve N absorption and transport rates through the regulation of key nitrogen transporter genes, thereby enhancing N utilization efficiency and promoting plant growth. Synopsis: EMF could enhance the efficiency of N utilization and promote the growth of Pinus massoniana under conditions of increased N deposition. [ABSTRACT FROM AUTHOR]

Enzyme behaviour is characterised in the laboratory using diluted solutions of enzyme. However, in vivo processes usually occur at [ST] ≈ [ET] ≈ Km. Furthermore, the study of enzyme action involves characterisation of inhibitors and their mechanisms. However, to date, there have been no reports proposing mathematical expressions that can be used to describe enzyme activity at high enzyme concentration apart from the simplest single substrate, irreversible case. Using a continued fraction approach, equations can be easily derived for the most common cases in monosubstrate reactions, such as irreversible or reversible reactions and effector (inhibitor or activator) kinetic interactions. These expressions are an extension of the classical Michaelis-Menten equations. A first analysis using these expressions permits to deduce some differences at high versus low enzyme concentration, such as the greater effectiveness of allosteric inhibitors compared to catalytic ones. Also, they can be used to understand catalyst saturation in a reaction. Although they can be linearised, these equations also show differences that need to be taken into account. For example, the different meaning of line intersection points in Dixon plots. All in all, these expressions may be useful tools for modelling in vivo and biotechnological processes. [ABSTRACT FROM AUTHOR]

Enzymes have several excellent catalytic features, and the last few years have seen a revolution in biocatalysis, which has grown from using one enzyme to using multiple enzymes in cascade reactions, where the product of one enzyme reaction is the substrate for the subsequent one. However, enzyme stability remains an issue despite the many benefits of using enzymes in a catalytic system. When enzymes are exposed to harsh process conditions, deactivation occurs, which changes the activity of the enzyme, leading to an increase in reaction time to achieve a given conversion. Immobilization is a well-known strategy to improve many enzyme properties, if the immobilization is properly designed and controlled. Enzyme co-immobilization is a further step in the complexity of preparing a biocatalyst, whereby two or more enzymes are immobilized on the same particle or support. One crucial problem when designing and using co-immobilized enzymes is the possibility of using enzymes with very different stabilities. This paper discusses different scenarios using two co-immobilized enzymes of the same or differing stability. The effect on operational performance is shown via simple simulations using Michaelis–Menten equations to describe kinetics integrated with a deactivation term. Finally, some strategies for overcoming some of these problems are discussed. [ABSTRACT FROM AUTHOR]

Poly(ethylene terephthalate) (PET) is one of the main synthetic plastics produced worldwide. The extensive use of this polymer causes several problems due to its low degradability. In this scenario, biocatalysts dawn as an alternative to enhance PET recycling. The enzymatic hydrolysis of PET results in a mixture of terephthalic acid (TPA), ethylene glycol (EG), mono-(2-hydroxyethyl) terephthalate (MHET) and bis-(2-hydroxyethyl) terephthalate (BHET) as main products. This work developed a new methodology to quantify the hydrolytic activity of biocatalysts, using BHET as a model substrate. The protocol can be used in screening enzymes for PET depolymerization reactions, amongst other applications. The very good fitting (R2 = 0.993) between experimental data and the mathematical model confirmed the feasibility of the Michaelis-Menten equation to analyze the effect of BHET concentration (8–200 mmol L−1) on initial hydrolysis rate catalyzed by Humicola insolens cutinase (HiC). In addition to evaluating the effects of enzyme and substrate concentration on the enzymatic hydrolysis of BHET, a novel and straightforward method for MHET synthesis was developed using an enzyme load of 0.025 g protein g BHET −1 and BHET concentration of 60 mmol L−1 at 40 °C. MHET was synthesized with high selectivity (97 %) and yield (82 %). The synthesized MHET properties were studied using differential scanning calorimetry (DSC), thermogravimetry (TGA), and proton nuclear magnetic resonance (1H NMR), observing the high purity of the final product (86.7 %). As MHET is not available commercially, this synthesis using substrate and enzyme from open suppliers adds new perspectives to monitoring PET hydrolysis reactions. • A novel and straightforward method for MHET synthesis was developed with high selectivity and yield. • The synthesized MHET properties were studied using differential analytical techniques. • Methodology to quantify the hydrolytic activity of biocatalysts was developed using BHET to understand the enzyme selectivity for PET hydrolysis. [ABSTRACT FROM AUTHOR]

The modern definition of enzymology is synonymous with the Michaelis–Menten equation instituted by Leonor Michaelis and Maud Menten. Most textbooks, or chapters within, discussing enzymology start with the derivation of the equation under the assumption of rapid equilibrium (as done by Michaelis–Menten) or steady state (as modified by Briggs and Haldane) conditions to highlight the importance of this equation as the bedrock on which interpretation of enzyme kinetic results is dependent. However, few textbooks or monographs take the effort of placing the equation within its right historical context and discuss the assumptions that have gone into its institution. This guide will dwell on these in substantial detail. Further, this guide will attempt to instil a sense of appreciation for the mathematical curve rectangular hyperbola, its unique attributes and how ubiquitous the curve is in biological systems. To conclude, this guide will discuss the limitations of the equation, and the method it embodies, and trace the journey of how investigators are attempting to move beyond the steady‐state approach and the Michaelis–Menten equation into full progress curve, pre–steady state and single‐turnover kinetic analysis to obtain greater insights into enzyme kinetics and catalysis. [ABSTRACT FROM AUTHOR]

The increasing interest in natural health-promoting compounds, which are mostly plant secondary metabolites, inspired attempts to stimulate mechanisms strengthening their bioaccumulation in crop plants via abiotic stress while maintaining the yield potential. This study investigates the long-term effects of limiting nitrogen (N) supply on the concentration of total phenolics, free radical activity of natural antioxidants, betacyanin content, biomass production, net photosynthetic rate, total chlorophyll content, and plant water relations in red beetroot plants (Beta vulgaris L.) grown hydroponically. Depending on fertilization, the range of N supply for evaporative demand comprises two contrasted nutrient zones, in which N is limiting (zone-1) or non-limiting (zone-2). Based on the carbon-nutrient-balance hypothesis, at the transition from 1st-zone to 2nd-zone, there is a narrow transition zone in which the plant nutrient status is considered 'critical'. Herein, to determine the 'critical' zone, a modified Michaelis-Menten (M-M) model was used using a piecewise linear regression on two indexes: net photosynthetic rates and free radical-scavenging capacity of phenolic antioxidants. The model showed that the 'critical' transition points of net photosynthetic rate and phenolic free radical content are located in a narrow zone ranging between 196.70 ± 8.75 and 271.54 ± 75.50 ppm NO 3 −, while the cropping season appears to affect slightly the range of 'critical' (transition) zone. Thus, supplying N to red beetroot plants to levels ranging within this 'critical' zone may be an efficient, profitable and sustainable way to increase the accumulation of health-promoting plant bioactive compounds (total phenolic compounds with radical activity and betacyanins) in hydroponically cultivated reed beetroot plants. [Display omitted] • Between primary and secondary metabolism there is a narrow 'critical' zone. • Low rates of N fertilization stimulate biosynthesis of bioactive compounds. • N supply within a 'critical' zone increases sustainability of plant production systems. [ABSTRACT FROM AUTHOR]

The kinetics of solute transport shed light on the roles these processes play in cellular physiology, and the absolute values of the kinetic parameters that quantitatively describe transport are increasingly used to model its impact on drug clearance. However, accurate assessment of transport kinetics is challenging. Although most carrier-mediated transport is adequately described by the Michaelis-Menten equation, its use presupposes that the rates of uptake used in the analysis of maximal rates of transport (Jmax) and half-saturation constants (Kt) reflect true unidirectional rates of influx from known concentrations of substrate. Most experimental protocols estimate the initial rate of transport from net substrate accumulation determined at a single time point (typically between 0.5 and 5 min) and assume it reflects unidirectional influx. However, this approach generally results in systematic underestimates of Jmax and overestimates of Kt; the former primarily due to the unaccounted impact of efflux of accumulated substrate, and the latter due to the influence of unstirred water layers. Here, we describe the bases of these timedependent effects and introduce a computational model that analyzes the time course of net substrate uptake at several concentrations to calculate Jmax and Kt for unidirectional influx, taking into account the influence of unstirred water layers and mediated efflux. This method was then applied to calculate the kinetics of transport of 1-methyl-4-phenylpryridinium and metformin by renal organic cation transporter 2 as expressed in cultured Chinese hamster ovary cells. NEW & NOTEWORTHY Here, we describe a mathematical model that uses the time course of net substrate uptake into cells from several increasing concentrations to calculate unique kinetic parameters [maximal rates of transport (Jmax) and half-saturation constants (Kt)] of the process. The method is the first to take into consideration the common complicating factors of unstirred layers and carrier-mediated efflux in the experimental determination of transport kinetics. [ABSTRACT FROM AUTHOR]

The research on inorganic nanozymes remains very active since the first paper on the "Intrinsic peroxidase-like properties of ferromagnetic nanoparticles" was published in Nature Nanotechnology in 2007. However, there is still the question, "What are the inorganic nanozymes?" Inorganic nanomaterials with intrinsic enzyme-like activities were not generated from artificial synthesis processing. The unique architecture of the metal site in inorganic nanomaterials contributes to the specificity and catalytic behavior described by the classic Michaelis–Menten equation, just like an enzyme (protein) or ribozyme (RNA), and as supported by density functional theory calculations. The discovered inorganic nanozymes participate in many key biological catalysis processes, especially related to oxygen. Possibly, they were essential to the emergence of life before RNAs and proteins, and they still play a crucial role in our daily lives, especially from the view of biogeochemistry. These inorganic nanozymes are classified as "inorganic enzymes", in line with the existing classifications of enzymes (proteins) and ribozymes (RNAs) as biocatalysts. By drawing inspiration from the metal architecture in nanomaterials, scientists can design artificial biocatalysts or functionalized nanozymes that are more stable and specific. It is necessary to pursue research to utilize inorganic nanozymes in the current biomedical applications and comprehend how they impact our Earth's environment. [ABSTRACT FROM AUTHOR]

Designing sustainable efficient peroxidase nanozymes and elucidating its peroxidase-like mechanism are very important for constructing colorimetric sensing platforms. Herein, a sustainable peroxidase nanozyme, porphyrin functionalized montmorillonite (PorMMT) loaded Cu 2 O-Fe 3 O 4 nanocomposites, is prepared by the two-step hydrothermal method and can be recycled conveniently by virtue of magnetism of Fe 3 O 4 from the composite (Cu 2 O-Fe 3 O 4 /PorMMT). Cu 2 O-Fe 3 O 4 /PorMMT peroxidase nanozyme can quickly catalyze the oxidation of the colourless substrate 3,3′,5,5′-tetramethylbiphenylamine (TMB) by H 2 O 2 into a blue product (oxTMB) only in 3 min. Compared with pure MMT, Cu 2 O and Fe 3 O 4 individual, Cu 2 O-Fe 3 O 4 /PorMMT exhibits the higher peroxidase-like activity. The catalytic activity of Cu 2 O-Fe 3 O 4 /PorMMT is reached the maximum at pH = 4 and T = 50 ℃. The catalytic behaviors of Cu 2 O-Fe 3 O 4 /PorMMT follow the Michaelis-Menten kinetic equation. The catalytic mechanism of Cu 2 O-Fe 3 O 4 /PorMMT is from the produced active species including superoxide free radicals (•O 2 −) and holes (h+). Based on the peroxidase-like activity of Cu 2 O-Fe 3 O 4 /PorMMT, a fast convenient colorimetric sensing platform has been designed to detect H 2 O 2 and hydroquinone (HQ), respectively. H 2 O 2 detection can be realized in the linear range of 5 10 mM with the limit of detection (LOD) of 2.68 mM, as well as hydroquinone (HQ) in the linear range of 5 50 μM with a much lower LOD of 4.37 μM. The convenient visual sensing platform has a potential application in monitoring HQ remains in lakes and rivers near dressing plants. [Display omitted] [ABSTRACT FROM AUTHOR]

[Display omitted] • The enzymatic activity was enhanced by 28.73 % with low intensity ultrasound (LU). • Amino acids obtained from sludge protein via UEH was optimized. • UEH was accurately modeled with Michaelis-Menten and protein hydrolysis model. • Enhanced enzymatic activity by LU was probably via altering enzyme conformation. • The nutrition and safety of sludge amino acids were confirmed. The recovery of high-value products such as complex amino acids from activated sludge is a crucial alternative for the recycling of excess sludge, although several challenges still need to be addressed, especially in terms of extraction efficiency and product quality. Herein, a promising approach involving low-intensity ultrasound-assisted enzymatic hydrolysis (UEH) was utilized to obtain complex amino acids from sludge protein. Compared with the values obtained for single enzymatic hydrolysis (EH), after a 20-minute pre-treatment at ambient temperature using low-intensity ultrasound (0.15 W/mL), the enzyme activity increased by 28.7 %, and the degree of protein hydrolysis (DH) correspondingly rose by 93.3 %. The application of ultrasound during subsequent EH resulted in a decrease in DH, with the extent of the decrease increasing over time. This suggested that the primary cause was conformational changes of enzymes rather than mass transfer. Moreover, both the Michaelis–Menten model (R2 > 0.99) and protein hydrolysis model (R2 > 0.96) accurately simulated EH and UEH, indicating that UEH exhibits superior enzyme–substrate affinity and a higher rate of protein hydrolysis. Furthermore, after separation and purification, the total content of seven essential amino acids in the dry material exceeded that in traditional feed sources such as soybean meal. Additionally, the safety test and energy consumption analysis demonstrated that UEH has good potential for the extraction of complex amino acids from excess sludge. [ABSTRACT FROM AUTHOR]

The British Journal of Clinical Pharmacology's April issue focuses on pharmacokinetics, with articles covering both scientific and applied aspects of drug development. The importance of understanding the fundamental properties of medicines is emphasized, and the use of artificial intelligence and modeling in exploring dose regimens and predicting responses is highlighted. Specific articles in this issue discuss renal and hepatic drug elimination, alterations in metabolic enzymes and their impact on medicine absorption and metabolism, and a less invasive method for phenotyping cytochrome P450 3A. Additionally, the issue includes studies on atrial fibrillation and anti-Shiga toxin fragments, as well as other topics such as drug use during pregnancy and lactation, pharmacological interventions for SARS-CoV-2, drug use in pediatrics, and the emulation of randomized trials. [Extracted from the article]

Natural enzymes are special proteins with excellent catalytic activity and selectivity. It is difficult to purify, store without losing the catalytic ability. Therefore, it is of great significance to study mimetic enzymes to realize the functions of natural enzymes and overcome their shortcoming. Herein, lentinan was used as a biological template to prepare palladium nanoparticles stabilized by lentinan (Pd‐LNT NPs), whose structure, activity and function were characterized. In addition, the 3,3′,5,5′‐tetramethylbenzidine (TMB) was oxidized by Pd‐LNT NPs in the presence of H2O2, which proved that they had good peroxidase‐like activity. The optimum reaction conditions were 30 °C and pH 4, the catalytic kinetic parameters were consistent with Michaelis‐Menten equation, and the good catalytic performance was verified. Furthermore, the colorimetric reaction based on the good peroxidase‐like activity of Pd‐LNT NPs was used for the convenient detection of H2O2. The linear range of colorimetric detection of H2O2 was 3.571‐107.143 μM, and it is worth noting that the detection limit was low to 4.714 μM. At last, Pd‐LNT NPs as biomimetic enzymes were used to detect the H2O2 released from HeLa cells, indicating they have great potential in the field of biological application. [ABSTRACT FROM AUTHOR]

[Display omitted] • Bi 2 Fe 4 O 9 nanozyme as a simple and low-cost colorimetric sensor was developed for easy dopamine detection. • The sensor achieved good performance with a linear range from 0.15 to 50 μM and a detection limit of 51 nM. • The nanozyme was reused up to 4 cycles without a significant decrease in its performance. This work describes the preparation of ternary bismuth ferrite oxide nanoparticles (Bi 2 Fe 4 O 9 NPs) with an enzyme mimetic activity for dopamine (DA) qualitative and quantitative detection. Bi 2 Fe 4 O 9 NPs were prepared using a facile, low cost, and one-pot hydrothermal treatment. The chemical composition, morphology, and optical properties of Bi 2 Fe 4 O 9 nanozyme were characterized using different techniques such as Fourier-transform infrared spectroscopy (FTIR), X-ray diffraction pattern (XRD), X-ray photoelectron spectroscopy (XPS), thermo-gravimetric analysis (TGA), dynamic light scattering (DLS), field-emission scanning electron microscopy (FESEM) imaging, FESEM-energy dispersive X-ray spectroscopy (EDS), UV–vis absorption, and fluorescence emission spectroscopy. Bi 2 Fe 4 O 9 NPs were utilized to catalyze the oxidation of a typical chromogenic peroxidase substrate, 3,3′,5,5′-tetramethylbenzidine (TMB), to form the blue-colored oxidized product (oxTMB), in the presence of hydrogen peroxide (H 2 O 2). All reactions occurred in acetate buffer solution (pH 3.5) to generate hydroxyl radicals (•OH) and the kinetics were followed by UV–vis absorbance at 654 nm. The steady-state kinetic parameters were obtained from the Michaelis-Menten equation and exhibited a good catalytic efficiency of Bi 2 Fe 4 O 9 NPs as enzyme mimetics. Michaelis–Menten constant (K m) values were estimated as 0.07 and 0.73 mM for TMB and H 2 O 2 , respectively. The presented method is efficient, rapid, cost-effective, and sensitive for the colorimetric detection of dopamine with a linear range (LR) from 0.15 to 50 μM and a detection limit (LOD) of 51 nM. The proposed colorimetric sensor was successfully applied for the detection of different concentrations of dopamine in spiked fetal bovine serum (FBS) and horse serum (HS) samples. It is anticipated that Bi 2 Fe 4 O 9 nanozyme holds great potential in biomedical analysis and diagnostic applications of dopamine-related diseases. [ABSTRACT FROM AUTHOR]

The area where a progress curve exhibitsmaximum curvature contains the most information about kinetic parameters. To determine these parameters more accurately from progress curves, we propose an iterative approach that calculates the area of maximum curvature based on an estimate of kinetic parameters and then recalculates the parameters based on time-concentration data points within this area. Based on this algorithm, we developed a computer script called iFIT as a free web application at http://www.i-fit.si. The benefits of working with iFIT are that it decreases the importance of initial substrate concentration and the impact of certain side reactions on the final calculated kinetic parameters. [ABSTRACT FROM AUTHOR]

The role of dimer formation for the onset of catalytic activity of SARS-CoV-2 main protease (MProWT) was assessed using a predominantly monomeric mutant (MProM). Rates of MProWT and MProM catalyzed hydrolyses display substrate saturation kinetics and second-order dependency on the protein concentration. The addition of the prodrug GC376, an inhibitor of MProWT, to MProM leads to an increase in the dimer population and catalytic activity with increasing inhibitor concentration. The activity reaches a maximum corresponding to a dimer population in which one active site is occupied by the inhibitor and the other is available for catalytic activity. This phase is followed by a decrease in catalytic activity due to the inhibitor competing with the substrate. Detailed kinetics and equilibrium analyses are presented and a modified Michaelis-Menten equation accounts for the results. These observations provide conclusive evidence that dimer formation is coupled to catalytic activity represented by two equivalent active sites. The binding of a drug targeting the active site of a predominantly monomeric SARS-CoV-2 main protease (MProM) favors an equilibrium shift to MProM dimer formation with two equivalent active sites. These results suggest targeting the monomeric active site and/or the dimer interface to interfere with the conformational rearrangements to active dimer formation as an alternative drug design strategy against MPro. [ABSTRACT FROM AUTHOR]

Enzymes produced by roots and soil microorganisms have been the biological driving factors of organic matter decomposition to nutrient cycling in soil. However, the enzyme investigation remains a technical question, due to the heterogeneity of spatial distribution around the roots in the rhizosphere. Soil in-situ zymogram can directly display the distribution enzyme activity by fluorescence mapping. This study aims to investigate the spatial distribution of soil enzyme activities in the rhizosphere of spring maize with different soil surface mulching in Weibei dryland. Taking the spring maize “Xianyu 335” as the research object, a field experiment was carried out without damaging the crop roots by soil in-situ zymogram in June 2020 at the Loess Plateau agroecological experimental station of the Chinese Academy of Sciences. A complete randomized block design was employed with three treatments of Straw Mulching (SM), Plastic Film Mulching (FM), and no-mulching (CK). The in-situ zymography of the root profile was obtained at the silking stage of spring maize. The reaction parameters were calculated with the Michaelis-Menten equation after the enzyme kinetic experiment and simulation. Besides, the correlations between the concentration of enzyme and the soil chemical/physical properties variables were determined for the driving factors of enzyme spatial distribution with the different mulching treatments. The results show that there was the maximum distribution area of β- Glucosidase Activity (BG) in the rhizosphere of SM treatment, which was 1.9 times that of plastic film mulching, and 8.1 times that of CK treatment. The longest distance (2.5 mm) along the roots of βglucosidase distribution was found with the SM treatment, which was 0.7 mm further than that with the FM, and 1.5mm further than that with CK. Correspondingly, the kinetic fitting showed that there were the maximum enzyme kinetic parameters Vmax and km of SM treatment, which was the fastest turnover time of soil substrate. The total distribution area of Leucine Aminopeptidase Activity (LAP) with the FM was the largest, which was 1.8 times that of SM, and 6.4 times that of CK. The longest distribution distance of enzyme activity with FM treatment was 4.5 mm, 1.7 and 2.1 mm further than those with the SM, and CK treatment, respectively. There were the maximum enzyme kinetic parameters with the FM treatment, all the same as the turnover of the substrate. The CK treatment presented the smallest hot spot area and kinetic parameters and the longest substrate turnover time, compared with the rest. The SM treatment increased the distribution area and extent of BG activity, indicating the fastest turnover rate of soil substrate, using the input of C, water storage, and soil N contents improvement. The FM treatment increased the distribution area and extent of LAP activity, with the fastest turnover rate of soil substrate using the soil temperature and water improvement. The CK treatment presented the smallest distribution area and extent of BG and LAP activities, where there was no change in the C input and soil temperature. As such, an enzyme kinetic mechanism was achieved to regulate the spatial distribution of enzyme activity in the crop rhizosphere soil for more nutrient uptake, particularly for the high yield and high nutrients utilization efficiency of dryland maize. [ABSTRACT FROM AUTHOR]

The kinetically derived maximal dose (KMD) provides a toxicologically relevant upper range for the determination of chemical safety. Here, we describe a new way of calculating the KMD that is based on sound Bayesian, theoretical, biochemical, and toxicokinetic principles, that avoids the problems of relying upon the area under the curve (AUC) approach that has often been used. Our new, mathematically rigorous approach is based on converting toxicokinetic data to the overall, or system-wide, Michaelis–Menten curve (which is the slope function for the toxicokinetic data) using Bayesian methods and using the "kneedle" algorithm to find the "knee" or "elbow"—the point at which there is diminishing returns in the velocity of the Michaelis–Menten curve (or acceleration of the toxicokinetic curve). Our work fundamentally reshapes the KMD methodology, placing it within the well-established Michaelis–Menten theoretical framework by defining the KMD as the point where the kinetic rate approximates the Michaelis–Menten asymptote at higher concentrations. By putting the KMD within the Michaelis–Menten framework, we leverage existing biochemical and pharmacological concepts such as "saturation" to establish the region where the KMD is likely to exist. The advantage of defining KMD as a region, rather than as an inflection point along the curve, is that a region reflects uncertainty and clarifies that there is no single point where the curve is expected to "break;" rather, there is a region where the curve begins to taper off as it approaches the asymptote (Vmax in the Michaelis–Menten equation). [ABSTRACT FROM AUTHOR]

The declining production of commercially important eucheumatoids related to serious problems, like increasing susceptibility to ice–ice disease and epiphytism, may be ameliorated by nutrition. This ushered an increasing interest in incorporating seaweeds into an integrated multi‐trophic aquaculture (IMTA) setup to take up excess inorganic nutrients produced by fish farms for their nourishment. In this regard, it is important to understand the nutrient uptake capacity of candidate seaweeds for incorporation into an IMTA system. Here, we examined the growth, nitrate (NO3‐) uptake kinetics, and biofiltration potential of Eucheuma denticulatum and three strains of Kappaphycus alvarezii (G‐O2, TR‐C16, and SW‐13) with distinct thallus morphologies. The NO3‐ uptake rates of the samples were determined under a range of NO3‐ concentrations (1–48 µM) and uptake rates were fitted to the Michaelis–Menten saturation equation. Among the examined eucheumatoids, only SW‐13 had a linear response to NO3‐ concentration while other strains had uptake rates that followed the Michaelis–Menten saturation equation. Eucheumadenticulatum had the lowest Km (9.78 ± 1.48 µM) while G‐O2 had the highest Vmax (307 ± 79.3 µmol · g−1 · min−1). The efficiency in NO3‐ uptake (highest Vmax/Km and α) was translated into the highest growth rate (3.41 ± 0.58% · d−1) measured in E. denticulatum. Our study provided evidence that eucheumatoids could potentially take up large amount of NO3‐ and fix CO2 when cultivated proximate to a fish farm as one component of an IMTA system. During a 45 ‐d cultivation period of eucheumatoids, as much as 370 g NO3‐ can be sequestered by every 1 kg initial biomass of E. denticulatum growing at 3% · d−1. Furthermore, based on our unpublished photosynthetic measurements, the congeneric K. striatus can fix 27.5 g C · kg−1 DW during a 12 h daylight period. [ABSTRACT FROM AUTHOR]

Introduction: Industrially, the use of enzymes has increased over time, as their production processes, stabilization and operating conditions have been optimized. Meanwhile associated with their use are minimized with environmental sustainability. The use of mathematical modeling has been a key factor in this advance, making it an great scientific interest area. Objective: To know the state of the art in this area of knowledge, highlighting the most successful and promising findings at present. Materials and Methods: A systematic search of recent high-impact publications between 2015 and 2021 was carried out in internationally recognized databases. A qualitative/comparative analysis of the approaches and contributions of the selected literature was carried out. Results and Discussion: It was evidenced that most of the mathematical models was developed from the Michaelis-Menten kinetic equation, but the simplifications assumed, the operating conditions and the mathematical methods used to solve these systems define the singularity and complexity of each investigation, which makes complex their comparison or implementation in a standardized way. Conclusions: In all cases, the proposed models were adjusted to experimental results or they agreed when different mathematical resolution methods were used. These results can provide valuable information for processes improvement and for equipment design, minimizing costs and time of experimentation and being more friendly to the environment. [ABSTRACT FROM AUTHOR]

Clonorchis sinensis (Cs) is a common trematode in Asian countries. Infection by Cs can result in many clinical symptoms. Here, a cDNA encoding a Cs apical sodium-dependent bile acid transporter (CsSBAT) was isolated from a Cs cDNA library, and functional characterization was performed using Xenopus laevis oocyte expression system. When expressed in Xenopus laevis oocytes, CsSBAT mediated the transport of radiolabeled estrone sulfate and dehydroepiandrosterone sulfate. No trans-uptake of carnitine, estradiol 17 β-D glucuronide, prostaglandin E2, p-aminohippuric acid, α-ketoglutaric acid, and tetraethylammonium was observed. CsSBAT-mediated estrone sulfate uptake was in a time- and sodium-dependent manner. CsSBAT showed no exchange properties in efflux experiments. Concentration-dependent results showed saturable kinetics consistent with the Michaelis–Menten equation. Nonlinear regression analyses yielded a Km value of 0.3 ± 0.04 μM for [3H]estrone sulfate. CsSBAT-mediated estrone sulfate uptake was strongly inhibited by sulfate conjugates but not glucuronide conjugates. These findings contribute to our understanding of CsSBAT transport properties and the cascade of estrogen metabolite movement in Cs. [ABSTRACT FROM AUTHOR]

Herein, the effect of inorganic carbon (IC) limitation on the nitrogen removal performance of the single-stage reactor containing nitritation and anammox gel carriers was investigated. As a result of a continuous feeding test, the effluent ammonium concentration increased as the IC concentration decreased, indicating the deterioration of nitritation activity, not anammox. Furthermore, the sensitivity of IC to anammox and nitritation activity was investigated in anammox and nitritation reactors, respectively. Consequently, the relationship between the effluent IC concentration and nitritation rate was well described using the Michaelis–Menten equation. The apparent K m value of nitritation was calculated as 4.4 mg-C L−1. In anammox reactor, it was calculated as 1.7 mg-C L−1. These results revealed that the affinity of nitritation gel carriers to IC was lower than that of anammox, supporting that nitritation activity was easily deactivated by decrease in the IC concentration rather than anammox. Microbial community analysis revealed that Nitrosomonas europaea and Candidatus Jettenia asiatica were the dominant species of ammonium-oxidizing and anammox bacteria. [Display omitted] • The effect of IC on the nitrogen removal performance of SSR was investigated. • The nitrogen removal performance was decreased with decrease of IC. • The effects of IC on nitritation and anammox were determined. • The affinity to IC of nitritation gel carriers was lower than that of anammox. • Dominant species in gel carriers were Nitrosomonas europaea and Candidatus Jettenia asiatica. [ABSTRACT FROM AUTHOR]

There is a vast amount of literature concerning the appropriateness of various perturbation parameters for the standard quasi-steady state approximation in the Michaelis–Menten reaction mechanism, and also concerning the relevance of these parameters for the accuracy of the approximation by the familiar Michaelis–Menten equation. Typically, the arguments in the literature are based on (heuristic) timescale estimates, from which one cannot obtain reliable quantitative estimates for the error of the quasi-steady state approximation. We take a different approach. By combining phase plane analysis with differential inequalities, we derive sharp explicit upper and lower estimates for the duration of the initial transient and substrate depletion during this transitory phase. In addition, we obtain rigorous bounds on the accuracy of the standard quasi-steady state approximation in the slow dynamics regime. Notably, under the assumption that the quasi-steady state approximation is valid over the entire time course of the reaction, our error estimate is of order one in the Segel–Slemrod parameter. [ABSTRACT FROM AUTHOR]

We study the temporal fluctuations in catalytic rates for single enzyme reactions undergoing slow transitions between two active states. We use a first passage time distribution formalism to obtain the closed-form analytical expressions of the mean reaction time and the randomness parameter for reaction schemes where conformational fluctuations are present between two free enzyme conformers. Our studies confirm that the sole presence of free enzyme fluctuations yields a non Michaelis-Menten equation and can lead to dynamic cooperativity. The randomness parameter, which is a measure of the dynamic disorder in the system, converges to unity at a high substrate concentration. If slow fluctuations are present between the enzyme-substrate conformers (off-pathway mechanism), dynamic disorder is present at a high substrate concentration. Our results confirm that the dynamic disorder at a high substrate concentration is determined only by the slowfluctuations between the enzyme-substrate conformers and the randomness parameter is greater than unity. Slow conformational fluctuations between free enzymes are responsible for the emergence of dynamic cooperativity in single enzymes. Our theoretical findings are well supported by comparison with experimental data on the single enzyme beta-galactosidase. [ABSTRACT FROM AUTHOR]

Depending on the chemical energy from ATP hydrolysis, myosin V can drive the multistep and continuous coupled cycling process to transport cellular cargo to targeted regions. However, it is still obscure how the molecular memory induced by the multistep coupled transported process could regulate the dynamic behavior of the motor state of myosin V. Here, we propose a novel non-Markovian polymorphic mechanochemical model to investigate the effect of the molecular memory on the mechanic of noise attenuation of myosin V system. We first define an effective transition rate for a multistep coupled reaction process which is the function of memory and system states to transform equivalently the non-Markovian process into the classical Markov process. By noise decomposition technology, it is observed that both the intrinsic and extrinsic noises of the ADP-myosin V bound state (AM ⋅ ADP) exhibit a monotonically decreasing trend with lengthening the molecular memory. Molecular memory as a regulation factor can amplify the contribution of intrinsic noise to the overall noise while reducing the influence of extrinsic noise on the AM ⋅ ADP. Moreover, the modulation of molecular memory could induce stochastic focusing. These results indicate that the role of molecular memory in the myosin V state transition can not only offer a handle to maintain the robustness of the motion system but also serve as a paradigm for studying more complex molecular motors. • Both the intrinsic and extrinsic noises of the ADP-myosin V bound state (AM ⋅ ADP) exhibit a monotonically decreasing trend with lengthening the molecular memory. • Molecular memory as a regulation factor can amplify the contribution of intrinsic noise. • Memory can induce to reduction of the influence of extrinsic noise on the AM ⋅ ADP. • The modulation of molecular memory could induce stochastic focusing. [ABSTRACT FROM AUTHOR]

Foodborne pathogenic bacteria are widespread in various foods, whose cross-contamination and re-contamination are critical influences on food safety. Rapid, accurate, and sensitive detection of foodborne pathogenic bacteria remains a topic of concern. CRISPR/Cas12a can recognize double-stranded DNA directly, showing great potential in nucleic acid detection. However, few studies have investigated the cleavage properties of CRISPR/Cas12a. In this study, the trans -cleavage properties of LbCas12a and AsCas12a were investigated to construct the detection methods for foodborne pathogenic bacteria. The highly sensitive fluorescent strategies for foodborne pathogens were constructed by analyzing the cleavage rates and properties of substrates at different substrate concentrations. Cas12a was activated in the presence of foodborne pathogenic target sequence was present, resulting in the cleavage of a single-stranded reporter ssDNA co-labelled by fluorescein quencher and fluorescein. The sensitivity and specificity of the Cas12a fluorescent strategy was investigated with Salmonella and Staphylococcus aureus as examples. The results showed that AsCas12a was slightly more capable of trans -cleavage than LbCas12a. The detection limits of AsCas12a for Salmonella and Staphylococcus aureus were 24.9 CFU mL−1 and 1.50 CFU mL−1, respectively. In all the seven bacteria, Staphylococcus aureus and Salmonella were accurately discriminated. The study provided a basis for constructing and improving the CRISPR/Cas12a fluorescence strategies. The AsCas12a-based detection strategy is expected to be a promising method for field detection. [Display omitted] • The trans -cleavage ability of LbCas12a and AsCas12a was compared. • AsCas12a showed better affinity and greater reaction rate with trans -cleaved substrates. • CRISPR/Cas12a accurately distinguished Salmonella and Staphylococcus aureus from five other food-borne pathogens. • AsCas12a fluorescence sensing strategy with good detection performance in food matrices. [ABSTRACT FROM AUTHOR]

Sensitive detection of cholesterol in animal-derived foods is crucial for maintaining human healthy diets. In this study, an elegant approach utilizing inorganic nanozyme-based magnetic mesoporous silica nanoparticles (MMSNs) for the highly sensitive detection of cholesterol in animal-derived food products was reported. The results revealed the fabricated MMSNs exhibited remarkably intrinsic peroxidase (POD)-like catalytic activities with improved affinity, and the catalytic behavior aligned well with Michaelis–Menten equation. In addition, the data indicated that the MMSNs enabled visual colorimetric detection of cholesterol with a remarkably low detection limit of 7.12 μM by combining catalytic oxidation with cholesterol oxidase (ChOx). Furthermore, the prepared MMSNs were successfully employed for assessing cholesterol content in milk and egg yolk samples, indicating potential applications for cholesterol detection in animal-derived foods in future. • An elegant inorganic nanozyme was proposed for cholesterol detection in animal-derived foods. • The fabricated MMSN nanozyme exhibited remarkably intrinsic peroxidase-like catalytic activities. • The MMSN nanozyme could sensitively determine cholesterol content in milk and egg yolk samples. • The MMSN nanozyme has great prospects for cholesterol detection in animal-derived foods in future. [ABSTRACT FROM AUTHOR]

Nanozymes were the novel research field to replace natural enzymes because of stability and low cost. However, the research on nanozymes was mainly focused on peroxidase, and there was little research about nanozymes with oxidase-like activity, especially mimic oxidase of small molecules related to human physiology. High levels of uric acid (UA) in the body can cause hyperuricemia and gout. And natural uricase cured this disease because it could oxidize UA. The oxidase-like activity of mixed valence state metal organic frameworks with cerium (MVSM) had been studied, but MVSM was found to have uricase-like activity in this article. The catalytic process of UA with MVSM was studied by a variety of analytical methods, which was similar to the natural uricase except for further oxidation of H2O2. The catalytic activity constants of MVSM were acquired by the Michaelis-Menten equation. MVSM had a better ability to catalyze UA in in vivo and in vitro experiments. An enzyme-free analysis–based mimic uricase for UA was established. All the experimental results proved that MVSM had a good prospect to replace the natural uricase. [ABSTRACT FROM AUTHOR]

The site selectivity for lysine conjugation on a native protein is difficult to control and characterize. Here, we applied mass spectrometry to examine the conjugation kinetics of Trastuzumab-IgG (Her-IgG) and α-lactalbumin under excess linker concentration ([L]0) based on the modified Michaelis–Menten equation, in which the initial rate constant per amine (kNH2 = Vmax/NH2/KM) was determined by the maximum reaction rate (Vmax/NH2) under saturated accessible sites and initial amine–linker affinity (1/KM). Reductive amination (RA) displayed 3–4 times greater Vmax/NH2 and a different panel of conjugation sites than that observed for N-hydroxysuccinimide ester (NHS) chemistry using the same length of polyethylene glycol (PEG) linkers. Moreover, faster conversion power rendered RA site selectivity among accessible amine groups and a greater tunable range of linker/protein ratio for aldehyde-linkers compared to those of the same length of NHS-linkers. Single conjugation with high yield or poly-conjugations with site homogeneity was demonstrated by controlling [L]0 or gradual addition to minimize the [L]0/KM ratio. Formaldehyde, the shortest aldehyde-linker with the greatest 1/KM, exhibited the highest selectivity and was shown to be a suitable probe to predict conjugation profile of aldehyde-linkers. Four linkers on the few probe-predicted hot spots were elucidated by kinetically controlled RA with conserved drug efficacy when conjugated with the payload. This study provides insights into controlling factors for homogenous and predictable amine bioconjugation. [ABSTRACT FROM AUTHOR]

The general characteristics of the effect of surfactants on the activity of lysozyme were demonstrated. The kinetics of bacterial cell lysis is consistent with the Michaelis-Menten equation and the presence of surfactants does not shift the pH-optimum of activity. Surfactants do not change the Km value but instead, affect the Vmax value. The experimental dependencies are well described by theoretical equations, which assume three surfactant binding sites on the lysozyme molecule. The dependencies of the activity of lysozyme on the surfactant concentration are either a step type (i.e., a higher plateau becomes a lower plateau), or a dependency with a maximum and continuation of the curve in the form of a plateau but with an increase in the surfactant concentration. It can be assumed that there is a mechanism for the regulation of lysozyme activity by an unknown natural factor that has a suitable hydrophobic radical capable of binding to the surface of lysozyme. [Display omitted] • The dependencies of the activity of lysozyme on various surfactants have common features. • The experimental data are well described by the theoretical curve, assuming three ligand binding sites by lysozyme. • It is possible that the activity of lysozyme can be regulated by natural factors that have suitable hydrophobic radicals. [ABSTRACT FROM AUTHOR]

Biochemical systems consist of numerous elementary reactions governed by the law of mass action. However, experimentally characterizing all the elementary reactions is nearly impossible. Thus, over a century, their deterministic models that typically contain rapid reversible bindings have been simplified with non-elementary reaction functions (e.g., Michaelis-Menten and Morrison equations). Although the non-elementary reaction functions are derived by applying the quasi-steady-state approximation (QSSA) to deterministic systems, they have also been widely used to derive propensities for stochastic simulations due to computational efficiency and simplicity. However, the validity condition for this heuristic approach has not been identified even for the reversible binding between molecules, such as protein-DNA, enzyme-substrate, and receptor-ligand, which is the basis for living cells. Here, we find that the non-elementary propensities based on the deterministic total QSSA can accurately capture the stochastic dynamics of the reversible binding in general. However, serious errors occur when reactant molecules with similar levels tightly bind, unlike deterministic systems. In that case, the non-elementary propensities distort the stochastic dynamics of a bistable switch in the cell cycle and an oscillator in the circadian clock. Accordingly, we derive alternative non-elementary propensities with the stochastic low-state QSSA, developed in this study. This provides a universally valid framework for simplifying multiscale stochastic biochemical systems with rapid reversible bindings, critical for efficient stochastic simulations of cell signaling and gene regulation. To facilitate the framework, we provide a user-friendly open-source computational package, ASSISTER, that automatically performs the present framework. Author summary: As experimentally characterizing all underlying processes of reactions in biochemical systems is almost impossible, their combined effects have frequently been described by simplified non-elementary reaction functions (e.g., Hill and Morrison functions). Recently, the deterministically driven non-elementary reaction functions have been heuristically used for stochastic simulations with the Gillespie algorithm. While this approach has been one of the most popular methods for efficient stochastic simulations, its accuracy has been controversial. Here, we finally solve this enigmatic open question. Specifically, we derive a complete condition under which this approach can accurately capture the stochastic dynamics of reversible binding, the critical reaction to describe nearly all biochemical systems such as gene regulation and enzyme catalysis. We illustrate that the use of this approach outside the identified range of validity seriously distorts the stochastic dynamics of a bistable switch in the cell cycle and an oscillator in the circadian clock. Importantly, to overcome this inaccuracy, we propose alternative simplified reaction functions for stochastic reversible binding. Combining the existing and proposed reaction functions, we have developed a computational package, ASSISTER, that performs universally valid stochastic model reduction. This enables accurate and efficient stochastic simulations of multiscale biochemical systems with rapid reversible bindings under any conditions. [ABSTRACT FROM AUTHOR]

This paper discusses a mathematical model of the diffusion and reaction in amperometric biosensor response with immobilized enzyme electrodes within a uniform film. This model contains a nonlinear differential equation related to Michaelis-Menten kinetics. In this paper, shifted Legendre wavelet method (SLWM) has been applied to obtain the mediator and substrate concentrations. The efficiency of the method is confirmed by means of the computational run time. The obtained approximate solutions are compared with the available results. Satisfactory agreement with homotopy perturbation method (HPM) and relaxation method (RM) is observed. Moreover, the shifted Legendre wavelet method is found to be simple, efficient and computationally attractive. [ABSTRACT FROM AUTHOR]

To understand the behaviour of complex systems, it is often necessary to use models that describe the dynamics of subnetworks. It has previously been established using projection methods that such subnetwork dynamics generically involves memory of the past and that the memory functions can be calculated explicitly for biochemical reaction networks made up of unary and binary reactions. However, many established network models involve also Michaelis-Menten kinetics, to describe, e.g., enzymatic reactions. We show that the projection approach to subnetwork dynamics can be extended to such networks, thus significantly broadening its range of applicability. To derive the extension, we construct a larger network that represents enzymes and enzyme complexes explicitly, obtain the projected equations, and finally take the limit of fast enzyme reactions that gives back Michaelis-Menten kinetics. The crucial point is that this limit can be taken in closed form. The outcome is a simple procedure that allows one to obtain a description of subnetwork dynamics, including memory functions, starting directly from any given network of unary, binary, and Michaelis-Menten reactions. Numerical tests show that this closed form enzyme elimination gives a much more accurate description of the subnetwork dynamics than the simpler method that represents enzymes explicitly and is also more efficient computationally. [ABSTRACT FROM AUTHOR]

A facile and green approach to the preparation of peroxidase-like nanozymes by reducing and functionalizing graphene oxide (rGO) with Ganoderma polysaccharide (GP) has been achieved in this work. Our results showed that the as-fabricated nanozyme, namely rGO-GP, possessed the excellent property of simulating peroxidase with higher catalytic activity compared with GO or rGO obtained by using chitosan, which may be due to the better dispersion of rGO-GP in the solution. Steady-state kinetics studies further showed that the catalytic process conformed to Michaelis-Menten equation and ping-pong mechanism. Benefiting from the excellent peroxidase property of rGO-GP, we have also successfully established a highly sensitive and selective colorimetric detection approach to trace detection of l-cysteine (l-Cys). The limit of detection (LOD) of l-cysteine is 0.1 μM and the linear detection range is 2–30 μM. Furthermore, the nanozyme was successfully applied for detecting l-cysteine in serum. This work therefore demonstrates the advantages of rGO-GP as an effective nanozyme in both its green synthesis and detecting application. [ABSTRACT FROM AUTHOR]

Urchin-like peroxidase mimics 5,10,15,20-tetrakis (4-carboxyl phenyl) porphyrin–functionalized CuCo2O4 nanospheres (Por-CuCo2O4) has been fabricated as an excellent visual biosensor. X-ray diffractometry (XRD), scanning electron microscopy (SEM), and X-ray photoelectron spectroscopy (XPS) have been employed to characterize the composition, morphologies, and elemental analysis of the as-synthesized Por-CuCo2O4. The catalytic activity of Por-CuCo2O4 was evaluated by the chromogenic substrate 3,3′,5,5′-tetramethylbenzidine (TMB) with the aid of H2O2, which exhibited a visual blue change with an absorption maximum at 652 nm for only 10 s. The peroxidase-like behaviors of Por-CuCo2O4 conformed to the Michaelis-Menten equation. Electrochemistry, radical scavenger, and fluorescence probe experiments verified that electron transfer, •O2− radicals, and holes (h+) are the important factors during the catalytic oxidation of TMB. Based on the inhibition of dopamine (DA) on TMB oxidation, the Por-CuCo2O4-based colorimetric biosensor has been successfully constructed for sensitive determination of DA witha detection limit (LOD) of 0.94 μΜ. In addition, colorimetry was validated to detect DA in serum samples with high sensitivity and good selectivity. [ABSTRACT FROM AUTHOR]