Topography of histone H3-H4 interaction with the Hat1-Hat2 acetyltransferase complex.

Chaperones influence histone conformation and intermolecular interaction in multiprotein complexes, and the structures obtained with full-length histones often provide more accurate and comprehensive views. Here, our structure of the Hat1-Hat2 acetyltransferase complex bound to Asf1-H3-H4 shows that the core domains of H3 and H4 are involved in binding Hat1 and Hat2, and the N-terminal tail of H3 makes extensive interaction with Hat2. These findings expand the knowledge about histone-protein interaction and implicate a function of Hat2/RbAp46/48, which is a versatile histone chaperone found in many chromatin-associated complexes, in the passing of histones between chaperones.

Allosteric communication between ligand binding domains modulates substrate inhibition in adenylate kinase.

Enzymes play a vital role in life processes; they control chemical reactions and allow functional cycles to be synchronized. Many enzymes harness large-scale motions of their domains to achieve tremendous catalytic prowess and high selectivity for specific substrates. One outstanding example is provided by the three-domain enzyme adenylate kinase (AK), which catalyzes phosphotransfer between ATP to AMP. Here we study the phenomenon of substrate inhibition by AMP and its correlation with domain motions. Using single-molecule FRET spectroscopy, we show that AMP does not block access to the ATP binding site, neither by competitive binding to the ATP cognate site nor by directly closing the LID domain. Instead, inhibitory concentrations of AMP lead to a faster and more cooperative domain closure by ATP, leading in turn to an increased population of the closed state. The effect of AMP binding can be modulated through mutations throughout the structure of the enzyme, as shown by the screening of an extensive AK mutant library. The mutation of multiple conserved residues reduces substrate inhibition, suggesting that substrate inhibition is an evolutionary well conserved feature in AK. Combining these insights, we developed a model that explains the complex activity of AK, particularly substrate inhibition, based on the experimentally observed opening and closing rates. Notably, the model indicates that the catalytic power is affected by the microsecond balance between the open and closed states of the enzyme. Our findings highlight the crucial role of protein motions in enzymatic activity.

Gibberellin-biosynthetic ent-kaurene synthases in higher plants do not require their non-catalytic domains for the catalysis.

ent-Kaurene is a biosynthetic intermediate diterpene of phytohormone gibberellins, and is biosynthesized from geranylgeranyl diphosphate via ent-copalyl diphosphate (ent-CDP). The successive cyclization is catalyzed by two distinct diterpene synthases, ent-CDP synthase (ent-CPS) and ent-kaurene synthase (KS). Homologs of these diterpene synthase genes have been reported to be involved in the biosynthesis of specialized-metabolic diterpenoids for defense in several plant species, including rice (Oryza sativa). These diterpene synthases consist of three domains, αßγ domains. Active sites of ent-CPS exist at the interface of ß and γ domain, while those of KS are located within the α domain. We herein carried out domain-deletion experiments using several KSs and KS like enzymes (KSLs) to obtain insights into the roles of domains other than active-site domains. As previously reported in taxadiene synthase, deletion of γ or ßγ domains drastically decreased activities of specialized-metabolic OsKSL5, OsKSL8, OsKSL7 and OsKSL10 in O. sativa. However, unexpectedly, only α domains of several gibberellin-biosynthetic KSs, including OsKS1 in O. sativa, AtKS in Arabidopsis thaliana, TaKS in wheat (Triticum aestivum) and BdKS1 in Brachypodium distachyon, retained their original functions. Additionally, the specialized-metabolic OsKSL4, which is closely related to OsKS1, also functioned without its ßγ domains. Domain-swapping experiments showed that replacing ßγ domains in OsKSL7 with those from other KS/KSLs retained the OsKSL7 activity. Moreover, deletion of ßγ domains of bifunctional PpCPS/KS in moss (Physcomitrella patens) drastically impaired its KS-related activity. Thus, we demonstrate that monofunctional gibberellin-biosynthetic KSs are the unique diterpene synthases that retain their functions without ßγ domains.

Control of protein activity by photoinduced spin polarized charge reorganization.

Considerable electric fields are present within living cells, and the role of bioelectricity has been well established at the organismal level. Yet much remains to be learned about electric-field effects on protein function. Here, we use phototriggered charge injection from a site-specifically attached ruthenium photosensitizer to directly demonstrate the effect of dynamic charge redistribution within a protein. We find that binding of an antibody to phosphoglycerate kinase (PGK) is increased twofold under illumination. Remarkably, illumination is found to suppress the enzymatic activity of PGK by a factor as large as three. These responses are sensitive to the photosensitizer position on the protein. Surprisingly, left (but not right) circularly polarized light elicits these responses, indicating that the electrons involved in the observed dynamics are spin polarized, due to spin filtration by protein chiral structures. Our results directly establish the contribution of electrical polarization as an allosteric signal within proteins. Future experiments with phototriggered charge injection will allow delineation of charge rearrangement pathways within proteins and will further depict their effects on protein function.

Identification of hydrogen bonding network for proton transfer at the quinol oxidation site of Rhodobacter capsulatus cytochrome bc1.

Cytochrome bc1 catalyzes electron transfer from quinol (QH2) to cytochrome c in reactions coupled to proton translocation across the energy-conserving membrane. Energetic efficiency of the catalytic cycle is secured by a two-electron and two-proton bifurcation reaction leading to oxidation of QH2 and reduction of the Rieske cluster and heme bL. The proton paths associated with this reaction remain elusive. Here, we used site-directed mutagenesis and quantum mechanical calculations to analyze the contribution of protonable side chains located at the heme bL side of the QH2 oxidation site in Rhodobacter capsulatus cytochrome bc1. We observe that the proton path is effectively switched off when H276 and E295 are simultaneously mutated to the nonprotonable residues in the H276F/E295V double mutant. The two single mutants, H276F or E295V, are less efficient but still transfer protons at functionally relevant rates. Natural selection exposed two single mutations, N279S and M154T, that restored the functional proton transfers in H276F/E295V. Quantum mechanical calculations indicated that H276F/E295V traps the side chain of Y147 in a position distant from QH2, whereas either N279S or M154T induce local changes releasing Y147 from that position. This shortens the distance between the protonable groups of Y147 and D278 and/or increases mobility of the Y147 side chain, which makes Y147 efficient in transferring protons from QH2 toward D278 in H276F/E295V. Overall, our study identified an extended hydrogen bonding network, build up by E295, H276, D278, and Y147, involved in efficient proton removal from QH2 at the heme bL side of QH2 oxidation site.

Computer-assisted semi-rational design enhanced the enzymatic activity and protein stability of Proteinase K in calcium-free conditions.

Wild-type Proteinase K binds to two Ca2+ ions, which play an important role in regulating enzymaticactivity and maintaining protein stability. Therefore, a predetermined concentration of Ca2+ must be added during the use of Proteinase K, which increases its commercial cost. Herein, we addressed this challenge using a computational strategy to engineer a Proteinase K mutant that does not require Ca2+ and exhibits high enzymatic activity and protein stability. In the absence of Ca2+, the best mutant, MT24 (S17W-S176N-D260F), displayed an activity approximately 9.2-fold higher than that of wild-type Proteinase K. It also exhibited excellent protein stability, retaining 56.2 % of its enzymatic activity after storage at 4 °C for 5 days. The residual enzymatic activity was 65-fold higher than that of the wild-type Proteinase K under the same storage conditions. Structural analysis and molecular dynamics simulations suggest that the introduction of new hydrogen bond and π-π stacking at the Ca2+ binding sites due to the mutation may be the reasons for the increased enzymatic activity and stability of MT24.

Nitrogen application in pod zone improves yield and quality of two peanut cultivars by modulating nitrogen accumulation and metabolism.

Cultivated peanut (Arachis hypogaea L.) represents one of the most important oil and cash crops world-widely. Unlike many other legumes, peanuts absorb nitrogen through their underground pods. Despite this unique feature, the relationship between yield and nitrogen uptake within the pod zone remains poorly understood. In our pot experiment, we divided the underground peanut part into two zones-pod and root-and investigated the physiological and agronomic traits of two peanut cultivars, SH11 (large seeds, LS) and HY23 (small seeds, SS), at 10 (S1), 20 (S2), and 30 (S3) days after gynophores penetrated the soil, with nitrogen application in the pod zone. Results indicated that nitrogen application increased pod yield, kernel protein content, and nitrogen accumulation in plants. For both LS and SS peanut cultivars, optimal nitrogen content was 60 kg·hm- 2, leading to maximum yield. LS cultivar exhibited higher yield and nitrogen accumulation increases than SS cultivar. Nitrogen application up-regulated the expression of nitrogen metabolism-related genes in the pod, including nitrate reductase (NR), nitrite reductase (NIR), glutamine synthetase (GS), glutamate synthase (NADH-GOGAT), ATP binding cassette (ABC), and nitrate transporter (NRT2). Additionally, nitrogen application increased enzyme activity in the pod, including NR, GS, and GOGAT, consistent with gene expression levels. These nitrogen metabolism traits exhibited higher up-regulations in the large-seeded cultivar than in the small-seeded one and showed a significant correlation with yield in the large-seeded cultivar at S2 and S3. Our findings offer a scientific basis for the judicious application and efficient utilization of nitrogen fertilization in peanuts, laying the groundwork for further elucidating the molecular mechanisms of peanut nitrogen utilization.

Maize/soybean intercropping increases nutrient uptake, crop yield and modifies soil physio-chemical characteristics and enzymatic activities in the subtropical humid region based in Southwest China.

Intercropping, a widely adopted agricultural practice worldwide, aims to increase crop yield, enhance plant nutrient uptake, and optimize the utilization of natural resources, contributing to sustainable farming practices on a global scale. However, the underlying changes in soil physio-chemical characteristics and enzymatic activities, which contribute to crop yield and nutrient uptake in the intercropping systems are largely unknown. Consequently, a two-year (2021-2022) field experiment was conducted on the maize/soybean intercropping practices with/without nitrogen (N) fertilization (i.e., N0; 0 N kg ha-1 and N1; 225 N kg ha-1 for maize and 100 N kg ha-1 for soybean ) to know whether such cropping system can improve the nutrients uptake and crop yields, soil physio-chemical characteristics, and soil enzymes, which ultimately results in enhanced crop yield. The results revealed that maize intercropping treatments (i.e., N0MI and N1MI) had higher crop yield, biomass dry matter, and 1000-grain weight of maize than mono-cropping treatments (i.e., N0MM, and N1MM). Nonetheless, these parameters were optimized in N1MI treatments in both years. For instance, N1MI produced the maximum grain yield (10,105 and 11,705 kg ha-1), biomass dry matter (13,893 and 14,093 kg ha-1), and 1000-grain weight (420 and 449 g) of maize in the year 2021 and 2022, respectively. Conversely, soybean intercropping treatments (i.e., N0SI and N1SI) reduced such yield parameters for soybean. Also, the land equivalent ratio (LER) and land equivalent ratio for N fertilization (LERN) values were always greater than 1, showing the intercropping system's benefits in terms of yield and improved resource usage. Moreover, maize intercropping treatments (i.e., N0MI and N1MI) and soybean intercropping treatments (i.e., N0SI and N1SI) significantly (p < 0.05) enhanced the nutrient uptake (i.e., N, P, K, Ca, Fe, and Zn) of maize and soybean, however, these nutrients uptakes were more prominent in N1MI and N1SI treatments of maize and soybean, respectively in both years (2021 and 2022) compared with their mono-cropping treatments. Similarly, maize-soybean intercropping treatments (i.e., N0MSI and N1MSI) significantly (p < 0.05) improved the soil-based N, P, K, NH4, NO3, and soil organic matter, but, reduced the soil pH. Such maize-soybean intercropping treatments also improved the soil enzymatic activities such as protease (PT), sucrose (SC), acid phosphatase (AP), urease (UE), and catalase (CT) activities. This indicates that maize-soybean intercropping could potentially contribute to higher and better crop yield, enhanced plant nutrient uptake, improved soil nutrient pool, physio-chemical characteristics, and related soil enzymatic activities. Thus, preferring intercropping to mono-cropping could be a preferable choice for ecologically viable agricultural development.

Investigating the synergistic effects of biochar, trans-zeatin riboside, and Azospirillum brasilense on soil improvement and enzymatic activity in water-stressed wheat.

BACKGROUND: Water stress is a major danger to crop yield, hence new approaches to strengthen plant resilience must be developed. To lessen the negative effects of water stress on wheat plants, present study was arranged to investigate the role of synergistic effects of biochar, trans-zeatin riboside (t-ZR), and Azospirillum brasilense on soil improvement and enzymatic activity in water-stressed wheat. RESULTS: In a three-replication experiment comprising of four treatments (T0: Control, T1: Drought stress (DS), T2: DS + t-ZR with biochar, T3: DS + A. brasilense with biochar), we observed notable improvements in soil quality and enzymatic activities in water-stressed wheat plants with the application of t-ZR and A. brasilense with biochar. In drought stress, Treatment having the application of A. brasilense with biochar performs best as compared to the other and significant increased the enzymatic activities such as peroxidase (7.36%), catalase (8.53%), superoxide dismutase (6.01%), polyphenol oxidase (14.14%), and amylase (16.36%) in wheat plants. Different enzymatic activities showed different trends of results. Soil organic C, dissolved organic C, dissolved organic N also enhanced 29.46%, 8.59%, 22.70% respectively with the application of A. brasilense with biochar under drought stress condition. CONCLUSIONS: The synergistic action of A. brasilense and biochar creates an effective microbiological environment that supports essential plant physiological processes during drought stress. This enhancement is attributed to improved soil fertility and increased organic matter content, highlighting the potential of these novel strategies in mitigating water stress effects and enhancing crop resilience.

What happens to Bifidobacterium adolescentis and Bifidobacterium longum ssp. longum in an experimental environment with eukaryotic cells?

BACKGROUND: The impact of probiotic strains on host health is widely known. The available studies on the interaction between bacteria and the host are focused on the changes induced by bacteria in the host mainly. The studies determining the changes that occurred in the bacteria cells are in the minority. Within this paper, we determined what happens to the selected Bifidobacterium adolescentis and Bifidobacterium longum ssp. longum in an experimental environment with the intestinal epithelial layer. For this purpose, we tested the bacteria cells' viability, redox activity, membrane potential and enzymatic activity in different environments, including CaCo-2/HT-29 co-culture, cell culture medium, presence of inflammatory inductor (TNF-α) and oxygen. RESULTS: We indicated that the external milieu impacts the viability and vitality of bacteria. Bifidobacterium adolescentis decrease the size of the live population in the cell culture medium with and without TNF-α (p < 0.001 and p < 0.01 respectively). In contrast, Bifidobacterium longum ssp. longum significantly increased survivability in contact with the eukaryotic cells and cell culture medium (p < 0.001). Bifidobacterium adolescentis showed significant changes in membrane potential, which was decreased in the presence of eukaryotic cells (p < 0.01), eukaryotic cells in an inflammatory state (p < 0.01), cell culture medium (p < 0.01) and cell culture medium with TNF-α (p < 0.05). In contrast, Bifidobacterium longum ssp. longum did not modulate membrane potential. Instead, bacteria significantly decreased the redox activity in response to milieus such as eukaryotic cells presence, inflamed eukaryotic cells as well as the culture medium (p < 0.001). The redox activity was significantly different in the cells culture medium vs the presence of eukaryotic cells (p < 0.001). The ability to ß-galactosidase production was different for selected strains: Bifidobacterium longum ssp. longum indicated 91.5% of positive cells, whereas Bifidobacterium adolescentis 4.34% only. Both strains significantly reduced the enzyme production in contact with the eukaryotic milieu but not in the cell culture media. CONCLUSION: The environmental-induced changes may shape the probiotic properties of bacterial strains. It seems that the knowledge of the sensitivity of bacteria to the external environment may help to select the most promising probiotic strains, reduce research costs, and contribute to greater reproducibility of the obtained probiotic effects.

Plant-soil interactions alter nitrogen and phosphorus dynamics in an advancing subarctic treeline.

Treelines advance due to climate warming. The impacts of this vegetation shift on plant-soil nutrient cycling are still uncertain, yet highly relevant as nutrient availability stimulates tree growth. Here, we investigated nitrogen (N) and phosphorus (P) in plant and soil pools along two tundra-forest transects on Kola Peninsula, Russia, with a documented elevation shift of birch-dominated treeline by 70 m during the last 50 years. Results show that although total N and P stocks in the soil-plant system did not change with elevation, their distribution was significantly altered. With the transition from high-elevation tundra to low-elevation forest, P stocks in stones decreased, possibly reflecting enhanced weathering. In contrast, N and P stocks in plant biomass approximately tripled and available P and N in the soil increased fivefold toward the forest. This was paralleled by decreasing carbon (C)-to-nutrient ratios in foliage and litter, smaller C:N:P ratios in microbial biomass, and lower enzymatic activities related to N and P acquisition in forest soils. An incubation experiment further demonstrated manifold higher N and P net mineralization rates in litter and soil in forest compared to tundra, likely due to smaller C:N:P ratios in decomposing organic matter. Overall, our results show that forest expansion increases the mobilization of available nutrients through enhanced weathering and positive plant-soil feedback, with nutrient-rich forest litter releasing greater amounts of N and P upon decomposition. While the low N and P availability in tundra may retard treeline advances, its improvement toward the forest likely promotes tree growth and forest development.

Construction and Application of a Pepsin-Functionalized Covalent Organic Framework with Prominent Chiral Recognition Ability.

The stable Pepsin@covalent organic framework (Pepsin@COF) were constructed base on matching COF pore diameter to pepsin dimension. It exhibits excellent chiral recognition capabilities (e. e. % up to 62.63 %) and potential for enantioseparation. Furthermore, a positive correlation between the immobilized enzyme activity and chiral recognition was revealed, offering insights for the design of biocatalytic nanosystems in chiral separation.

Antifungal activity of copper oxide nanoparticles derived from Zizyphus spina leaf extract against Fusarium root rot disease in tomato plants.

Incorporating green chemistry concepts into nanotechnology is an important focus area in nanoscience. The demand for green metal oxide nanoparticle production has grown in recent years. The beneficial effects of using nanoparticles in agriculture have already been established. Here, we highlight some potential antifungal properties of Zizyphus spina leaf extract-derived copper oxide nanoparticles (CuO-Zs-NPs), produced with a spherical shape and defined a 13-30 nm particle size. Three different dosages of CuO-Zs-NPs were utilized and showed promising antifungal efficacy in vitro and in vivo against the selected fungal strain of F. solani causes tomato root rot disease, which was molecularly identified with accession number (OP824846). In vivo  results indicated that, for all CuO-Zs-NPs concentrations, a significant reduction in Fusarium root rot disease occurred between 72.0 to 88.6% compared to 80.5% disease severity in the infected control. Although treatments with either the chemical fungicide (Kocide 2000) showed a better disease reduction and incidence with (18.33% and 6.67%) values, respectively, than CuO-Zs-NPs at conc. 50 mg/l, however CuO-Zs-NPs at 250 mg/l conc. showed the highest disease reduction (9.17 ± 2.89%) and lowest disease incidence (4.17 ± 3.80%). On the other hand, CuO-Zs-NPs at varied values elevated the beneficial effects of tomato seedling vigor at the initial stages and plant growth development compared to either treatment with the commercial fungicide or Trichoderma Biocide. Additionally, CuO-Zs-NPs treatments introduced beneficial results for tomato seedling development, with a significant increase in chlorophyll pigments and enzymatic activity for CuO-Zs-NPs treatments. Additionally, treatment with low concentrations of CuO-Zs-NPs led to a rise in the number of mature pollen grains compared to the immature ones.  however the data showed that CuO-Zs-NPs have a unique antifungal mechanism against F. solani, they  subsequently imply that CuO-Zs-NPs might be a useful environmentally friendly controlling agent for the Fusarium root rot disease that affects tomato plants.

Biochemical and molecular-level effects of co-exposure to chlorpyrifos and lambda-cyhalothrin on the earthworm (Eisenia fetida).

Farmland soil organisms frequently encounter pesticide mixtures presented in their living environment. However, the underlying toxic mechanisms employed by soil animals to cope with such combined pollution have yet to be explored. This investigation aimed to reveal the changes in cellular and mRNA levels under chlorpyrifos (CPF) and lambda-cyhalothrin (LCT) co-exposures in earthworms (Eisenia fetida). Results exhibited that the combination of CPF and LCT triggered an acute synergistic influence on the animals. Most exposures resulted in significant alterations in the activities of total superoxide dismutase (T-SOD), copper/zinc superoxide dismutase (Cu/Zn-SOD), caspase 3, and carboxylesterase (CarE) compared to the basal level. Moreover, when exposed to chemical mixtures, the transcription levels of four genes [heat shock protein 70 (hsp70), gst, sod, and calreticulin (crt)] also displayed more pronounced changes compared with their individual exposures. These changes in determined parameters indicated the occurrence of oxidative stress, cell death, detoxification dysfunction, and endoplasmic reticulum damage after co-exposure to CPF and LCT in E. fetida. The comprehensive examination of mixture toxicities of CPF and LCT at different endpoints would help to understand the overall toxicity they cause to soil invertebrates. The augmented deleterious effect of these pesticides in a mixture suggested that mixture toxicity assessment was necessary for the safety evaluation and application of pesticide mixtures.

Effects of different microplastic types on soil physicochemical properties, enzyme activities, and bacterial communities.

Global concern continues to mount regarding the accumulation of microplastics (MPs) in soil. However, little is known about how various types of MPs influence the properties of soil ecosystems. Here, we evaluated the effects of six different types of MPs, including low-density polyethylene (LDPE), polyamide (PA), polystyrene (PS), polyhydroxy-alkanoates (PHA), polybutadiene styrene (PBS), and polylactide (PLA), on soil physicochemical properties, enzyme activities, and microbial communities. At the end of a 230-day soil incubation, we observed significant changes in soil moisture content, soil organic carbon, pH, NH4+-N, NO3--N, and available phosphorus. The addition of MPs had a significant influence on the activities of soil ß-glucosidase, acid phosphatase, urease, and fluorescein diacetate hydrolase, with effects varying with MP type. Results of 16S rRNA gene high throughput sequencing showed that MP exposure had little effect on soil microbial alpha diversity, but that PHA contamination significantly reduced ACE, Chao1, and Shannon index values. MP contamination also altered soil microbial community composition. In particular, the relative abundance of Firmicutes increased significantly while the relative abundance of Actinobacteriota, Proteobacteria (especially the nitrogen-fixing rhizobia), and Acidobacteriota decreased following exposure to PHA. Redundancy analysis showed that acid phosphatase and pH were the two main environmental factors affecting bacterial community structure at the phylum and order levels. Furthermore, Tax4Fun2 analysis found that MP treatment disrupted fundamental bacterial metabolic pathways. Our findings indicate that different types of MPs can affect soil fertility, bacterial community structure, and function in various ways, and highlight that biodegradable MPs may alter soil bacterial communities more than conventional MPs.

Physiological evaluation of seasonal sperm quality in a biannual spawner, Pacific abalone: Effects on in-vitro fertilization and cryotolerance.

Pacific abalone, Haliotis discus hannai, is a highly valuable gastropod mollusk commonly found in Southeast Asia. The present study aims to analyze the seminal plasma quality, sperm quality, and cryotolerance of the Pacific abalone sperm during its reproductive season. The seminal plasma quality was evaluated by analyzing biochemical and metabolite composition, enzymatic activity (superoxide dismutase, catalase, and glutathione), and lipid peroxidation (LPO) activity. The sperm quality was evaluated by analyzing motility, concentration, volume, ATP content, acrosome integrity (AI), plasma membrane integrity (PMI), mitochondrial membrane potential (MMP), DNA integrity, and fertilization potential. The cryotolerance capacity was evaluated by analyzing post-thaw motility, AI, PMI, MMP, and DNA integrity. Seminal plasma osmolarity was significantly higher (1123.3 ± 1.5 mOsmL-1) in May compared to other reproductive periods, with Cl- (516.8 ± 0.5 mM) and Na+ (460.2 ± 0.4 mM) as the dominant ions. The seminal plasma pH remained constant at 6.8 throughout the reproductive season. Improved enzymatic activity and lower LPO were detected in May or June. Sperm quality indicators were similar in May and June, except for sperm production. The fertilization potential (May: 93.0 ± 4.4%, June: 86.0 ± 7.2%) and hatching rate (May: 86.6 ± 5.78%, June: 82.3 ± 3.2%) of Pacific abalone were significantly higher in May or June than they were in other reproductive seasons. The motility (May: 50.19 ± 2.35%, June: 49.96 ± 1.60%), AI (May: 44.02 ± 3.46%, June: 42.16 ± 3.61%), PMI (May: 54.12 ± 3.29%, June: 52.82 ± 2.58%), and MMP (May: 44.02 ± 3.46%, June: 42.16 ± 3.61%) of the cryopreserved sperm were similar in May and June compared with those preserved in other reproductive seasons. The DNA integrity of the cryopreserved sperm was similar in May (80.3 ± 6.7%) or June (78.9 ± 7.4%) and had a higher cryotolerance than in other reproductive seasons. Hence, it can be suggested that May and/or June are suitable periods for sperm physiology experiments, artificial reproduction, and sperm cryopreservation of Pacific abalone.

Identification of a novel nonsense mutation in α-galactosidase A that causes Fabry disease in a Chinese family.

Fabry disease, a lysosomal storage disease, is an uncommon X-linked recessive genetic disorder stemming from abnormalities in the alpha-galactosidase gene (GLA) that codes human alpha-Galactosidase A (α-Gal A). To date, over 800 GLA mutations have been found to cause Fabry disease (FD). Continued enhancement of the GLA mutation spectrum will contribute to a deeper recognition and underlying mechanisms of FD. In this study, a 27-year-old male proband exhibited a typical phenotype of Fabry disease. Subsequently, family screening for Fabry disease was conducted, and high-throughput sequencing was employed to identify the mutated gene. The three-level structure of the mutated protein was analyzed, and its subcellular localization and enzymatic activity were determined. Apoptosis was assessed in GLA mutant cell lines to confirm the functional effects. As a result, a new mutation, c.777_778del (p. Gly261Leufs*3), in the GLA gene was identified. The mutation caused a frameshift during translation and the premature appearance of a termination codon, which led to a partial deletion of the domain in C-terminal region and altered the protein's tertiary structure. In vitro experiments revealed a significant reduction of the enzymatic activity in mutant cells. The expression was noticeably decreased at the mRNA and protein levels in mutant cell lines. Additionally, the subcellular localization of α-Gal A changed from a homogeneous distribution to punctate aggregation in the cytoplasm. GLA mutant cells exhibited significantly higher levels of apoptosis compared to wild-type cells.

Human fecal alpha-glucosidase activity and its relationship with gut microbiota profiles and early stages of intestinal mucosa damage.

OBJECTIVES: We investigated potential relationships among initial lesions of the intestinal mucosa, fecal enzymatic activities and microbiota profiles. METHODS: Fecal samples from 54 volunteers were collected after recruitment among individuals participating in a colorectal cancer (CRC) screening program in our region (Northern Spain) or attending for consultation due to clinical symptoms; intestinal mucosa samples were resected during colonoscopy. Enzymatic activities were determined in fecal supernatants by a semi-quantitative method. The fecal microbiota composition was determined by 16S rRNA gene-based sequencing. The results were compared between samples from clinical diagnosis groups (controls and polyps), according with the type of polyp (hyperplastic polyps or conventional adenomas) and considering the grade of dysplasia for conventional adenomas (low and high grade dysplasia). RESULTS: High levels of α-glucosidase activity were more frequent among samples from individuals diagnosed with intestinal polyps, reaching statistical significance for conventional adenomas and for low grade dysplasia adenomas when compared to controls. Regarding the microbiota profiles, higher abundance of Christensenellaceae_R-7 group and Oscillospiraceae_UCG-002 were found in fecal samples displaying low α-glucosidase activity as compared with those with higher activity as well as in controls with respect to conventional adenomas. A relationship was evidenced among intestinal mucosal lesions, gut glucosidase activities and intestinal microbiota profiles. CONCLUSIONS: Our findings suggest a relationship among altered fecal α-glucosidase levels, the presence of intestinal mucosal lesions, which can be precursors of CRC, and shifts in defined microbial groups of the fecal microbiota.

Cypermethrin exposure: Implications for growth, phenotypic attributes, and biochemical responses in monosex tilapia (Oreochromis niloticus).

The present study was conducted to evaluate the effects of exposure to sublethal concentrations of cypermethrin on growth, phenotypic traits, and metabolic and antioxidant enzymes activities of monosex tilapia (Oreochromis niloticus). The median lethal concentration (LC50) of cypermethrin was initially ascertained to be 0.04 ppm after 12 h of exposure to three concentrations (0.04, 0.20, and 1.0 ppm) via the Probit analysis method. The sublethal exposures were obtained for 3 months at 0.0016, 0.008, and 0.04 ppm of cypermethrin. The growth performance, phenotypic traits, and enzymatic activity were analyzed. The research findings revealed a notable impact of cypermethrin on the growth performance of monosex tilapia, with a significant (p < 0.05) decrease in weight observed as the concentration increased. The experiment documented a significant change (p < 0.05) in the phenotypic traits of standard length, total length, tail length, trunk length, eye diameter, body area, and color patterns, but not dressing percentage. Furthermore, it was observed that the activities of amylase and protease in both the intestine and muscle decreased significantly (p < 0.05) with increasing concentrations of cypermethrin, except for amylase in the muscle, where no significant change was noted (p > 0.05). The results also demonstrated significant reductions (p < 0.05) in the activity of metabolic enzymes, specifically superoxide dismutase and catalase, following exposure to sublethal levels of cypermethrin. The study suggested that sublethal exposure of pyrethroid alters the growth, phenotypic traits, metabolic enzymes activities, and immunity of monosex tilapia.

Structural Characterization and Functional Analysis of Mevalonate Kinase from Tribolium castaneum (Red Flour Beetle).

Mevalonate kinase (MevK) is an important enzyme in the mevalonate pathway that catalyzes the phosphorylation of mevalonate into phosphomevalonate and is involved in juvenile hormone biosynthesis. Herein, we present a structure model of MevK from the red flour beetle Tribolium castaneum (TcMevK), which adopts a compact α/ß conformation that can be divided into two parts: an N-terminal domain and a C-terminal domain. A narrow, deep cavity accommodating the substrate and cofactor was observed at the junction between the two domains of TcMevK. Computational simulation combined with site-directed mutagenesis and biochemical analyses allowed us to define the binding mode of TcMevK to cofactors and substrates. Moreover, TcMevK showed optimal enzyme activity at pH 8.0 and an optimal temperature of 40 °C for mevalonate as the substrate. The expression profiles and RNA interference of TcMevK indicated its critical role in controlling juvenile hormone biosynthesis, as well as its participation in the production of other terpenoids in T. castaneum. These findings improve our understanding of the structural and biochemical features of insect Mevk and provide a structural basis for the design of MevK inhibitors.