Autoinhibition of ubiquitin-specific protease 8: insights into domain interactions and mechanisms of regulation.

Ubiquitin-specific proteases (USPs) are a family of multi-domain deubiquitinases (DUBs) with variable architectures, some containing regulatory auxiliary domains. Among the USP family, all occurrences of intramolecular regulation presently known are autoactivating. USP8 remains the sole exception as its putative WW-like domain, conserved only in vertebrate orthologs, is autoinhibitory. Here, we present a comprehensive structure-function analysis describing the autoinhibition of USP8 and provide evidence of the physical interaction between the WW-like and catalytic domains. The solution structure of full-length USP8 reveals an extended, monomeric conformation. Coupled with DUB assays, the WW-like domain is confirmed to be the minimal autoinhibitory unit. Strikingly, autoinhibition is only observed with the WW-like domain in cis and depends on the length of the linker tethering it to the catalytic domain. Modelling of the WW:CD complex structure and mutagenesis of interface residues suggests a novel binding site in the S1 pocket. To investigate the interplay between phosphorylation and USP8 autoinhibition, we identify AMP-activated protein kinase as a highly selective modifier of S718 in the 14-3-3 binding motif. We show that 14-3-3γ binding to phosphorylated USP8 potentiates autoinhibition in a WW-like domain-dependent manner by stabilizing an autoinhibited conformation. These findings provide mechanistic details on the autoregulation of USP8 and shed light on its evolutionary significance.

Cancer-associated mutations in protein kinase C theta are loss-of-function.

The Ca2+-independent, but diacylglycerol-regulated, novel protein kinase C (PKC) theta (θ) is highly expressed in hematopoietic cells where it participates in immune signaling and platelet function. Mounting evidence suggests that PKCθ may be involved in cancer, particularly blood cancers, breast cancer, and gastrointestinal stromal tumors, yet how to target this kinase (as an oncogene or as a tumor suppressor) has not been established. Here, we examine the effect of four cancer-associated mutations, R145H/C in the autoinhibitory pseudosubstrate, E161K in the regulatory C1A domain, and R635W in the regulatory C-terminal tail, on the cellular activity and stability of PKCθ. Live-cell imaging studies using the genetically-encoded fluorescence resonance energy transfer-based reporter for PKC activity, C kinase activity reporter 2 (CKAR2), revealed that the pseudosubstrate and C1A domain mutations impaired autoinhibition to increase basal signaling. This impaired autoinhibition resulted in decreased stability of the protein, consistent with the well-characterized behavior of Ca2+-regulated PKC isozymes wherein mutations that impair autoinhibition are paradoxically loss-of-function because the mutant protein is degraded. In marked contrast, the C-terminal tail mutation resulted in enhanced autoinhibition and enhanced stability. Thus, the examined mutations were loss-of-function by different mechanisms: mutations that impaired autoinhibition promoted the degradation of PKC, and those that enhanced autoinhibition stabilized an inactive PKC. Supporting a general loss-of-function of PKCθ in cancer, bioinformatics analysis revealed that protein levels of PKCθ are reduced in diverse cancers, including lung, renal, head and neck, and pancreatic. Our results reveal that PKCθ function is lost in cancer.

Stabilization of insect cell membranes and soluble enzymes by accumulated cryoprotectants during freezing stress.

Most multicellular organisms are freeze sensitive, but the ability to survive freezing of the extracellular fluids evolved in several vertebrate ectotherms, some plants, and many insects. Here, we test the coupled hypotheses that are perpetuated in the literature: that irreversible denaturation of proteins and loss of biological membrane integrity are two ultimate molecular mechanisms of freezing injury in freeze-sensitive insects and that seasonally accumulated small cryoprotective molecules (CPs) stabilize proteins and membranes against injury in freeze-tolerant insects. Using the drosophilid fly, Chymomyza costata, we show that seven different soluble enzymes exhibit no or only partial loss of activity upon lethal freezing stress applied in vivo to whole freeze-sensitive larvae. In contrast, the enzymes lost activity when extracted and frozen in vitro in a diluted buffer solution. This loss of activity was fully prevented by adding low concentrations of a wide array of different compounds to the buffer, including C. costata native CPs, other metabolites, bovine serum albumin (BSA), and even the biologically inert artificial compounds HistoDenz and Ficoll. Next, we show that fat body plasma membranes lose integrity when frozen in vivo in freeze-sensitive but not in freeze-tolerant larvae. Freezing fat body cells in vitro, however, resulted in loss of membrane integrity in both freeze-sensitive and freeze-tolerant larvae. Different additives showed widely different capacities to protect membrane integrity when added to in vitro freezing media. A complete rescue of membrane integrity in freeze-tolerant larvae was observed with a mixture of proline, trehalose, and BSA.

The crotonylated and succinylated proteins of jujube involved in phytoplasma-stress responses.

BACKGROUND: Protein posttranslational modifications (PTMs) are fast and early responses to environmental changes, including pathogen infection. Jujube witches' broom (JWB) is a phytoplasma disease causing great economic loss in jujube production. After phytoplasma infection, the transcriptional, translational, and metabolic levels in jujube were activated, enabling it to survive during phytoplasma invasion. However, no study has yet reported on PTMs in jujube. Lysine crotonylation (Kcr) and lysine succinylation (Ksu) have been popular studies in recent years and their function in plant phytoplasma-stress responses remains unclear. RESULTS: Here, 1656 crotonylated and 282 succinylated jujube proteins were first identified under phytoplasma-stress, of which 198 were simultaneously crotonylated and succinylated. Comparative analysis revealed that 656 proteins, 137 crotonylated and 43 succinylated proteins in jujube were regulated by phytoplasma infection, suggesting that Kcr was more universal than Ksu. Kcr differentially expressed proteins (DEPs) were related to ribosomes, photosynthetic and carbon metabolism, while Ksu DEPs were mainly involved in carbon metabolism, the TCA cycle and secondary metabolite biosynthesis. The crosstalk network among proteome, crotonylome and succinylome showed that DEPs related to ribosomal, peroxidases and glutathione redox were enriched. Among them, ZjPOD51 and ZjPHGPX2 significantly increased at the protein and Kcr level under phytoplasma-stress. Notably, 7 Kcr sites were identified in ZjPHGPX2, a unique antioxidant enzyme. After inhibitor nicotinamide (NAM) treatment, GPX enzyme activity in jujube seedlings was reduced. Further, site-directed mutagenesis of key Kcr modification sites K130 and/or K135 in ZjPHGPX2 significantly reduced its activity. CONCLUSIONS: This study firstly provided large-scale datasets of Kcr and Ksu in phytoplasma-infected jujube and revealed that Kcr modification in ZjPHGPX2 positively regulates its activity.

Adenylyl Cyclase Isoform 6 in the Pulmonary Artery Is Inhibited by Hypoxia via Cysteine Nitrosylation.

Persistent pulmonary hypertension of the newborn (PPHN) is a hypoxic disorder of pulmonary vascular relaxation, mediated in part by adenylyl cyclase (AC). Neonatal pulmonary arteries (PA) express mainly AC6 isoform, followed by AC3, 7 and 9. AC6 expression is upregulated in hypoxia. We reported AC enzyme inhibition due to S-nitrosylation in PPHN PA, and in PA myocytes exposed to hypoxia. We hypothesize that hypoxia promotes cysteine thiol nitrosylation of AC6, impairing cAMP production. HEK293T cells stably expressing AC isoforms (AC3, 5, 6, 7, 9), or cysteine-to-alanine mutants AC6_C1004A, AC6_C1145A or AC6_C447A were cultured in normoxia (21% O2) or hypoxia (10% O2) for 72 hours, or challenged with nitroso donor S-nitrosocysteine (CysNO). AC activity was determined by real-time live-cell cAMP measurement (cADDis assay) or terbium-norfloxacin AC catalytic assay, with or without challenge by allosteric agonist forskolin; protein S-nitrosylation detected by biotin switch method and quantified by affinity precipitation. Only AC6 catalytic activity is inhibited in hypoxia or by S-nitrosylating agent, in presence or absence of forskolin; impaired cAMP production in hypoxia correlates with increased cysteine nitrosylation of AC6. Selective AC6 inhibition in pulmonary artery myocytes extinguishes AC sensitivity to inhibition by hypoxia. Alanine substitution of C1004, but not of other cysteines, decreases S-nitrosylation of AC6. AC activity is diminished in AC6_C1004A compared to AC6 wild type. Substitution of C1004 also extinguishes the inhibition of AC6 by hypoxia. We conclude AC6 is uniquely S-nitrosylated in hypoxia, inhibiting its activity and cAMP generation. We speculate that S-nitrosylation at C1004 may inhibit AC6 interaction with Gαs, playing a role in PPHN pathophysiology.

The role of N-terminal phosphorylation of DGK-θ.

Diacylglycerol kinases (DGKs) are lipid kinases that mediate the phosphorylation of diacylglycerol (DAG) leading to the production of phosphatidic acid (PtdOH). To examine the role of phosphorylation on DGK-θ, we first identified the phosphorylated sites on endogenous DGK-θ from mouse brain and found four sites: S15, S17, which we refer to phosphomotif-1 sites, and S22 and S26 which we refer to as phosphomotif-2 sites. This study focused on the role of these phosphorylated sites on enzyme activity, membrane binding, thermal stability, and cellular half-life of DGK-θ. After generating a construct devoid of all non-catalytic phosphorylation sites (4A), we also generated other constructs to mimic phosphorylation of these residues by mutating them to glutamate (E). Our data demonstrate that an increase in membrane affinity requires the phosphorylation of all four endogenous sites as the phosphomimetic 4E but not other phosphomimietics. Furthermore, 4E also shows an increase in basal activity as well as an increase in the Syt1-induced activity compared to 4A. It is noteworthy that these phosphorylations had no effect on the thermal stability or cellular half-life of this enzyme. Interestingly, when only one phosphorylation domain (phosphomotif-1 or phosphomotif-2) contained phosphomimetics (S15E/S17E or S22E/S26E), the basal activity was also increased but membrane binding affinity was not increased. Furthermore, when only one residue in each domain mimicked an endogenous phosphorylated serine (S15E/S22E or S17E/S26E), the Syt1-induced activity as well as membrane binding affinity decreased relative to 4A. These results indicate that these endogenous phosphorylation sites contribute differentially to membrane binding and enzymatic activity.

Impact of Surfactants on Cumulative Trypsin Activity in Bottom-Up Proteome Analysis.

Trypsin digestion plays a pivotal role in successful bottom-up peptide characterization and quantitation. While denaturants are often incorporated to enhance protein solubility, surfactants are recognized to inhibit enzyme activity. However, several reports have suggested that incorporating surfactants or other solvent additives may enhance digestion and MS detection. Here, we assess the impacts of ionic surfactants on cumulative trypsin activity and subsequently evaluate the total digestion efficiency of a proteome mixture by quantitative MS. Although low surfactant concentrations, such as 0.01% SDS or 0.2% SDC, significantly enhanced the initial trypsin activity (by 14 or 42%, respectively), time course assays revealed accelerated enzyme deactivation, evident by 10- or 40-fold reductions in trypsin activity half-life at these respective surfactant concentrations. Despite enhanced initial tryptic activity, quantitative MS analysis of a common liver proteome extract, digested with various surfactants (0.01 or 0.1% SDS, 0.5% SDC), consistently revealed decreased peptide counts and signal intensity, indicative of a lower digestion efficiency compared to a nonsurfactant control. Furthermore, including detergents for digestion did not improve the detection of membrane proteins, nor hydrophobic peptides. These results stress the importance of assessing cumulative enzyme activity when optimizing the digestion of a proteome mixture, particularly in the presence of denaturants.

Additive impact of soy protein dietary intake and exercise on visceral fat mass reduction and mitochondrial complex I activation in skeletal muscle.

Soy protein has shown remarkable effectiveness in reducing fat mass compared with other protein sources, and exercise has the potential to further enhance this fat loss effect. Previous studies have demonstrated that soy protein intake leads to decreased fatty acid synthesis, which contributes to its fat-loss properties. However, the exact mechanism by which these lipids are consumed remains unclear. To investigate this, we conducted a comprehensive study using C57/BL6 male mice, comparing the effects of soy and casein proteins with and without exercise (Casein-Sed, Casein-Ex, Soy-Sed, and Soy-Ex groups) under high- and low-protein conditions (14% or 40% protein). Our findings revealed that combining soy protein intake with exercise significantly reduced epididymal white adipose tissue (eWAT) weight, particularly in the high-protein diet group. Further analysis revealed that exercise increased the expression of lipid oxidation-regulatory proteins, including mitochondrial oxidative phosphorylation protein (OXPHOS) complexes, in the plantaris muscle regardless of the protein source. Although soy protein intake did not directly affect muscle mitochondrial protein expression, the activity of OXPHOS complex I was additively enhanced by exercise and soy protein under the 40% protein condition. Notably, complex I activity inversely correlated with eWAT weight in the soy protein diet group. These results highlight the potential link between improved complex I activity induced by soy protein and fat mass reduction, which emphasizes the promising benefits of combining soy protein with exercise in promoting fat loss.NEW & NOTEWORTHY The findings revealed that soy protein intake combined with exercise resulted in reduced adipose tissue weight compared with that obtained with casein protein intake. Furthermore, the joint impact of exercise and soy protein consumption resulted in enhanced activity of oxidative phosphorylation protein (OXPHOS) complex I in fast-twitch muscles, which appears to be associated with fat mass reduction. These findings elucidate the potential additive effects of soy protein and exercise on body weight management.

Exploring the Genomic Landscape of Bacillus paranthracis PUMB_17 as a Proficient Phosphatidylcholine-Specific Phospholipase C Producer.

Phospholipases find versatile applications across industries, including detergent production, food modification, pharmaceuticals (especially in drug delivery systems), and cell signaling research. In this study, we present a strain of Bacillus paranthracis for the first time, demonstrating significant potential in the production of phosphatidylcholine-specific phospholipase C (PC-PLC). The investigation thoroughly examines the B. paranthracis PUMB_17 strain, focusing on the activity of PC-PLC and its purification process. Notably, the PUMB_17 strain displays extracellular PC-PLC production with high specific activity during the late exponential growth phase. To unravel the genetic makeup of PUMB_17, we employed nanopore-based whole-genome sequencing and subsequently conducted a detailed genome annotation. The genome comprises a solitary circular chromosome spanning 5,250,970 bp, featuring a guanine-cytosine ratio of 35.49. Additionally, two plasmids of sizes 64,250 bp and 5845 bp were identified. The annotation analysis reveals the presence of 5328 genes, encompassing 5186 protein-coding sequences, and 142 RNA genes, including 39 rRNAs, 103 tRNAs, and 5 ncRNAs. The aim of this study was to make a comprehensive genomic exploration that promises to enhance our understanding of the previously understudied and recently documented capabilities of Bacillus paranthracis and to shed light on a potential use of the strain in the industrial production of PC-PLC.

Succinimide Derivatives as Acetylcholinesterase Inhibitors-In Silico and In Vitro Studies.

We studied the effect of succinimide derivatives on acetylcholinesterase activity due to the interest in compounds that influence this enzyme's activity, which could help treat memory issues more effectively. The following parameters were established for this purpose based on kinetic investigations of the enzyme in the presence of succinimide derivatives: the half-maximal inhibitory concentration, the maximum rate, the inhibition constant, and the Michaelis-Menten constant. Furthermore, computational analyses were performed to determine the energy required for succinimide derivatives to dock with the enzyme's active site. The outcomes acquired in this manner demonstrated that all compounds inhibited acetylcholinesterase in a competitive manner. The values of the docking energy parameters corroborated the kinetic parameter values, which indicated discernible, albeit slight, variations in the inhibitory intensity among the various derivatives.

The correlation of asparaginase enzyme activity levels after PEG-asparaginase administration with clinical characteristics and adverse effects in Chinese paediatric patients with acute lymphoblastic leukaemia.

Studies on asparaginase enzyme activity (AEA) monitoring in Chinese patients receiving PEG-asparaginase remain limited. We monitored AEA in paediatric patients diagnosed with acute lymphoblastic leukaemia (ALL) and treated according to the Chinese Children's Cancer Group study protocols, CCCG-ALL-2015/CCCG-ALL-2020 protocols. We measured the AEA at days 7 ± 1 and 14 ± 1 and analysed their association with patient characteristics and PEG-asparaginase-related adverse effects (AEs). We measured 2147 samples from 329 patients. Mean AEA levels (interquartile range) were 931 iu/L (654-1174 iu/L) at day 7 ± 1 and 664 iu/L (463-860 iu/L) at day 14 ± 1. The AEA levels were higher in younger children and increased with the cumulative dose numbers. PEG-asparaginase inactivation rate was 19.1%, and the silent inactivation (SI) rate was 12.5%. Nine patients were identified with allergic-like reactions. Hypofibrinogenaemia, hypertriglyceridaemia, pancreatitis and thrombosis were associated with older age, whereas hypoglycaemia was associated with younger age. The risk of hypertriglyceridaemia and hypoglycaemia increased with cumulative dose numbers of PEG-asparaginase. Except for hypofibrinogenaemia, elevated AEA levels did not increase the risk of PEG-asparaginase-related AEs. Drug monitoring can be utilized as guidance for treatment decision-making. Individualizing asparaginase doses do not reduce toxicities. The treatment target of PEG-asparaginase remains to achieve sustained and adequate activity.

Semi-rational design based on the interaction between SmFMO and FAD isoalloxazine ring to enhance the enzyme activity.

Flavin monooxygenases (FMOs) have been widely used in the biosynthesis of natural compounds due to their excellent stereoselectivity, regioselectivity and chemoselectivity. Stenotrophomonas maltophilia flavin monooxygenase (SmFMO) has been reported to catalyze the oxidation of various thiols to corresponding sulfoxides, but its activity is relatively low. Herein, we obtained a mutant SmFMOF52G which showed 4.35-fold increase in kcat/Km (4.96 mM-1s-1) and 6.84-fold increase in enzyme activity (81.76 U/g) compared to the SmFMOWT (1.14 mM-1s-1 and 11.95 U/g) through semi-rational design guided by structural analysis and catalytic mechanism combined with high-throughput screening. By forming hydrogen bond with O4 atom of FAD isoalloxazine ring and reducing steric hindrance, the conformation of FAD isoalloxazine ring in SmFMOF52G is more stable, and NADPH and substrate are closer to FAD isoalloxazine ring, shortening the distances of hydrogen transfer and substrate oxygenation, thereby increasing the rate of reduction and oxidation reactions and enhancing enzyme activity. Additionally, the overall structural stability and substrate binding capacity of the SmFMOF52G have significant improved than that of SmFMOWT. The strategy used in this study to improve the enzyme activity of FMOs may have generality, providing important references for the rational and semi-rational engineering of FMOs.

Transcriptome profiling reveals the impact of various levels of biochar application on the growth of flue-cured tobacco plants.

BACKGROUND: Biochar, a carbon-rich source and natural growth stimulant, is usually produced by the pyrolysis of agricultural biomass. It is widely used to enhance plant growth, enzyme activity, and crop productivity. However, there are no conclusive studies on how different levels of biochar application influence these systems. METHODS AND RESULTS: The present study elucidated the dose-dependent effects of biochar application on the physiological performance, enzyme activity, and dry matter accumulation of tobacco plants via field experiments. In addition, transcriptome analysis was performed on 60-day-old (early growth stage) and 100-day-old (late growth stage) tobacco leaves to determine the changes in transcript levels at the molecular level under various biochar application levels (0, 600, and 1800 kg/ha). The results demonstrated that optimum biochar application enhances plant growth, regulates enzymatic activity, and promotes biomass accumulation in tobacco plants, while higher biochar doses had adverse effects. Furthermore, transcriptome analysis revealed a total of 6561 differentially expressed genes (DEGs) that were up- or down-regulated in the groupwise comparison under different treatments. KEGG pathways analysis demonstrated that carbon fixation in photosynthetic organisms (ko00710), photosynthesis (ko00195), and starch and sucrose metabolism (ko00500) pathways were significantly up-regulated under the optimal biochar dosage (600 kg/ha) and down-regulated under the higher biochar dosage (1800 kg/ha). CONCLUSION: Collectively, these results indicate that biochar application at an optimal rate (600 kg/ha) could positively affect photosynthesis and carbon fixation, which in turn increased the synthesis and accumulation of sucrose and starch, thus promoting the growth and dry matter accumulation of tobacco plants. However, a higher biochar dosage (1800 kg/ha) disturbs the crucial source-sink balance of organic compounds and inhibits the growth of tobacco plants.

Nano silica's role in regulating heavy metal uptake in Calendula officinalis.

BACKGROUND: Soil contamination with heavy metals poses a significant threat to plant health and human well-being. This study explores the potential of nano silica as a solution for mitigating heavy metal uptake in Calendula officinalis. RESULTS: Greenhouse experiments demonstrated, 1000 mg•kg- 1 nano silica caused a 6% increase in soil pH compared to the control treatment. Also in 1000 mg. kg- 1 nano silica, the concentrations of available Pb (lead), Zn (zinc), Cu (copper), Ni (nickel), and Cr (chromium) in soil decreased by 12%, 11%, 11.6%, 10%, and 9.5%, respectively, compared to the control. Nano silica application significantly reduces heavy metal accumulation in C. officinalis exposed to contaminated soil except Zn. In 1000 mg.kg- 1 nano silica shoots Zn 13.28% increased and roots Zn increased 13% compared to the control treatment. Applying nano silica leads to increase the amount of phosphorus (P) 25%, potassium (K) 26% uptake by plant, In 1000 mg.kg - 1 treatment the highest amount of urease enzyme activity was 2.5%, dehydrogenase enzyme activity, 23.6% and the highest level of alkaline phosphatase enzyme activity was 13.5% higher than the control treatment. CONCLUSION: Nano silica, particularly at a concentration of 1000 mg.kg - 1, enhanced roots and shoots length, dry weight, and soil enzyme activity Moreover, it increased P and K concentrations in plant tissues while decreasing heavy metals uptake by plant.

The impact of root systems and their exudates in different tree species on soil properties and microorganisms in a temperate forest ecosystem.

BACKGROUND: The species composition of tree stands plays an important role in shaping the properties of forest soils. The aim of our research was to determine the influence on soil properties of the root systems of six species of trees which form forest stands in the temperate climatic zone. The research covered areas including six tree species - Scots pine (Pinus sylvestris L.), European larch (Larix deciduas Mill.), English oak (Quercus robur L.), English ash (Fraxinus excelsior L.), European beech (Fagus sylvatica L.) and European hornbeam (Carpinus betulus L.). In our study, we determined the characteristics of the roots and the amount of carbon excreted alongside their exudates. Enzymatic activity, and the composition and diversity of the fungi and bacteria, were also determined in addition to the basic physicochemical properties of the soil samples. RESULTS: A strong relationship between the root characteristics and soil properties, including the pH, basic cation content and phosphorus content, was confirmed. In addition, the enzymatic activity of phosphatase, ß-glucosidase, N-acetyl-ß-D-glucosaminidase and ß-D-cellobiosidase were positively correlated with the root characteristics. The study on soil bacteria across different tree species revealed Proteobacteria and Actinobacteriota to be the most abundant phylum. Fungal analysis showed Basidiomycota and Ascomycota as the dominant phyla. Ascomycota dominated in hornbeam and oak soils. Mortierellomycota was remarkably more present in pine soil. CONCLUSIONS: This analysis of root systems and soil properties confirmed the distinctness of ash stands, which were also more abundant in various microorganisms. It was also found that soils affected by different tree species were characterised by varied fungal and bacterial composition. The ash had particularly beneficial impact on soil microbiota.

Confirming the enzymatic activity and neurodevelopmental trajectory of PYCR1 mutation in one child with autosomal-recessive cutis laxa type 2.

Autosomal-recessive cutis laxa type 2 (ARCL2) is a rare genetic disorder caused by pyrroline-5-carboxylate reductase 1 (PYCR1) mutations and characterized by loose and sagging skin, typical facial features, intrauterine growth retardation, and developmental delay. To study the effect of PYCR1 mutations on protein function and clinical features, we identified a homozygous missense mutation c.559G > A (p.Ala187Thr) in PYCR1 in a Chinese child with typical clinical features, especially severe developmental delays. The three-dimensional (3D) model showed the modification of the hydrogen bonds produce a misfolding in the mutant PYCR1 protein. Mutagenesis and enzyme assay study revealed decreased activity of the mutant protein in vitro, indicating that this mutation impairs PYCR1 function. Our findings confirmed abnormal enzymatic activity and neurodevelopmental trajectory of this PYCR1 mutation.

Multi-Enzyme Activity of MIL-101 (Fe)-Derived Cascade Nano-Enzymes for Antitumor and Antimicrobial Therapy.

The clinical application of oncology therapy is hampered by high glutathione concentrations, hypoxia, and inefficient activation of cell death mechanisms in cancer cells. In this study, Fe and Mo bimetallic sulfide nanomaterial (FeS2@MoS2) based on metal-organic framework structure is rationally prepared with peroxidase (POD)-, catalase (CAT)-, superoxide dismutase (SOD)-like activities and glutathione depletion ability, which can confer versatility for treating tumors and mending wounds. In the lesion area, FeS2@MoS2 with SOD-like activity can facilitate the transformation of superoxide anions (O2 -) to hydrogen peroxide (H2O2), and then the resulting H2O2 serves as a substrate for the Fenton reaction with FMS to produce highly toxic hydroxyl radicals (∙OH). Simultaneously, FeS2@MoS2 has an ability to deplete glutathione (GSH) and catalyze the decomposition of nicotinamide adenine dinucleotide phosphate (NADPH) to curb the regeneration of GSH from the source. Thus it can realize effective tumor elimination through synergistic apoptosis-ferroptosis strategy. Based on the alteration of the H2O2 system, free radical production, glutathione depletion and the alleviation of hypoxia in the tumor microenvironment, FeS2@MoS2 NPS can not only significantly inhibit tumors in vivo and in vitro, but also inhibit multidrug-resistant bacteria and hasten wound healing. It may open the door to the development of cascade nanoplatforms for effective tumor treatment and overcoming wound infection.

Rationally Designed Functionalization of Single-Walled Carbon Nanotubes for Real-Time Monitoring of Cholinesterase Activity and Inhibition in Plasma.

Enzymes play a pivotal role in regulating numerous bodily functions. Thus, there is a growing need for developing sensors enabling real-time monitoring of enzymatic activity and inhibition. The activity and inhibition of cholinesterase (CHE) enzymes in blood plasma are fluorometrically monitored using near-infrared (NIR) fluorescent single-walled carbon nanotubes (SWCNTs) as probes, strategically functionalized with myristoylcholine (MC)- the substrate of CHE. A significant decrease in the fluorescence intensity of MC-suspended SWCNTs upon interaction with CHE is observed, attributed to the hydrolysis of the MC corona phase of the SWCNTs by CHE. Complementary measurements for quantifying choline, the product of MC hydrolysis, reveal a correlation between the fluorescence intensity decrease and the amount of released choline, rendering the SWCNTs optical sensors with real-time feedback in the NIR biologically transparent spectral range. Moreover, when synthetic and naturally abundant inhibitors inhibit the CHE enzymes present in blood plasma, no significant modulations of the MC-SWCNT fluorescence are observed, allowing effective detection of CHE inhibition. The rationally designed SWCNT sensors platform for monitoring of enzymatic activity and inhibition in clinically relevant samples is envisioned to not only advance the field of clinical diagnostics but also deepen further understanding of enzyme-related processes in complex biological fluids.

Integrating Biocatalysts into Metal-Organic Frameworks: Disentangling the Roles of Affinity, Molecular Weight, and Size.

The integration of biocatalysts within metal-organic frameworks (MOFs) is attracting growing interest due to its potential to both enhance biocatalyst stability and sustain biocatalyst activity in organic solvents. However, the factors that facilitate the post-synthetic infiltration of such large molecules into MOF pores remain unclear. This systematic study enabled the identification of the influence of biocatalyst molecular size, molecular weight and affinity on the uptake by an archetypal MOF, NU-1000. We analyzed a range of six biocatalysts with molecular weights from 1.9 kDa to 44.4 kDa, respectively. By employing a combination of fluorescence tagging and 3D-STED confocal laser scanning microscopy, we distinguished between biocatalysts that were internalized within the MOF pores and those sterically excluded. The catalytic functions of the biocatalysts hosted within the MOF were investigated and found to show strong variations relative to the solvated case, ranging from a two-fold increase to a strong decrease.

How does CHD4 slide nucleosomes?

Chromatin remodelling enzymes reposition nucleosomes throughout the genome to regulate the rate of transcription and other processes. These enzymes have been studied intensively since the 1990s, and yet the mechanism by which they operate has only very recently come into focus, following advances in cryoelectron microscopy and single-molecule biophysics. CHD4 is an essential and ubiquitous chromatin remodelling enzyme that until recently has received less attention than remodellers such as Snf2 and CHD1. Here we review what recent work in the field has taught us about how CHD4 reshapes the genome. Cryoelectron microscopy and single-molecule studies demonstrate that CHD4 shares a central remodelling mechanism with most other chromatin remodellers. At the same time, differences between CHD4 and other chromatin remodellers result from the actions of auxiliary domains that regulate remodeller activity by for example: (1) making differential interactions with nucleosomal epitopes such as the acidic patch and the N-terminal tail of histone H4, and (2) inducing the formation of distinct multi-protein remodelling complexes (e.g. NuRD vs ChAHP). Thus, although we have learned much about remodeller activity, there is still clearly much more waiting to be revealed.