Aspartate Research Articles

Comparative Study on the Proximate Composition and Amino Acid Levels in Chanterelle (Cantharellus cibarius) and Wood Ear (Auricularia auricula) Mushroom Samples

Mushrooms have been consumed by mankind for millennia. They are macrofungi with a distinctive fruiting body which can be hypogeous or epigeous, large enough to be seen with unaided eyes. They are not widely consumed due to lack of information on its nutritive value. Therefore, this research was carried out to compare the proximate gross energy composition as well as amino acids levels in two selected edible mushrooms (Cantharellus cibarius and Auricularia auricula). Fully matured species of Chanterelle mushroom (Cantherellus cibarius) and Wood ear mushroom (Auricularia auricula) were collected from different parts of Southern and Northern Cross River; Ikom and Bekwara precisely. The collected samples were air dried for 5 days and then stored in transparent polythene bag for analysis. The dried and stored mushroom were pounded to powdered form and then subjected to amino acid analysis using isocratic HPLC and proximate analysis for proximate composition using the standard methods of Association of Official Analytical Chemist (AOAC). The results of this research showed that chanterelle mushroom has high concentration of amino acids compared to Wood ear mushroom. However, there was significant differences (p<0.05) in the levels of isoleucine in both mushrooms. There were no significant differences in the proximate composition and gross energy levels of both mushroom species. Some of the amino acids detected in both mushrooms were: methionine, threonine, isoleucine, leucine, phenylalanine, valine, tryptophan, histidine, arginine, aspartic acid, glycine, alanine and glutamic acid. Based on the findings of this study, the two edible mushrooms were observed to have appreciable levels of gross energy and amino acids. However, Chanterelle mushroom had higher protein composition than wood ear. Hence, it can be concluded that Chanterelle mushroom has greater nutritive value and more suitable for consumption.

Construction of Chiral Covalent Organic Frameworks Through a Linker Decomposition Chiral Induction Strategy for Circularly Polarized Light Detection

AbstractExploring new strategies for construction of chiral covalent organic frameworks (COFs) is of paramount importance yet remains a challenge. Herein, we report the rational design and construction of chiral COFs through a linker decomposition chiral induction (LDCI) strategy. Three pairs of azine‐linked chiral COFs are successfully synthesized by the condensation reactions of C3‐symmetric 4,4′,4′′‐(1,3,5‐triazine‐2,4,6‐triyl)tribenzaldehyde (Tz) with flexible chiral dihydrazide linkers derived from malic acid, aspartic acid and tartaric acid, respectively. Remarkably, upon complete or partial decomposition from flexible chiral dihydrazides to hydrazine during COF synthesis, the homochirality of these COFs, originating from the single‐handedness conformation of propeller‐like Tz cores, is well preserved. Such a stereoselective chiral memory realized via the LDCI strategy is confirmed by time‐dependent powder X‐ray diffraction (PXRD), Fourier transform infrared (FT‐IR) and diffuse reflectance circular dichroism (DRCD). Moreover, the resultant azine‐linked chiral COFs are used as the active materials to fabricate photodetectors to directly distinguish circularly polarized light (CPL), showing impressive recognition performances on the identification of left‐handed circularly (LHC) and right‐handed circularly (RHC) polarized lights. Notably, the residual undecomposed flexible chiral linkers within the COFs are found to be conducive to improving the polarization discrimination ratio. This work highlights LDCI as a new and effective strategy for constructing homochiral COFs with promising future in chiral optical application.

Bone-Targeted Fluoropeptide Nanoparticle Inhibits NF-κB Signaling to Treat Osteosarcoma and Tumor-Induced Bone Destruction.

Osteosarcoma is a malignant bone cancer usually characterized by symptoms of bone loss due to pathologically enhanced osteoclast activity. Activated osteoclasts enhance bone resorption and promote osteosarcoma cell progression by secreting various cytokines. Intercepting the detrimental interplay between osteoclasts and osteosarcoma cells is considered as an option for osteosarcoma treatment. Here, a bone-targeted fluoropeptide nanoparticle that can inhibit the nuclear factor kappa B (NF-κB) signaling in both osteoclasts and osteosarcoma to address the above issue is developed. The NF-κB essential modulator binding domain (NBD) peptide is conjugated with a fluorous tag to improve its proteolytic stability and intracellular penetration. The NBD peptide is efficiently delivered into cells after fluorination to induce apoptosis of osteocarcoma cells, and inhibits osteoclasts differentiation. The fluorous-tagged NBD peptide is further co-assembled with an oligo (aspartic acid) terminated fluoropeptide to form bone-targeted peptide nanoparticles for osteosarcoma treatment. The targeted nanoparticles efficiently inhibited tumor progression and osteosarcoma-induced bone destruction in vivo. This co-assembled fluoropeptide nanoplatform proposed in this study offers a promising approach for targeted and intracellular delivery of peptide therapeutics in the treatment of various diseases.

Construction of Chiral Covalent Organic Frameworks Through a Linker Decomposition Chiral Induction Strategy for Circularly Polarized Light Detection

AbstractExploring new strategies for construction of chiral covalent organic frameworks (COFs) is of paramount importance yet remains a challenge. Herein, we report the rational design and construction of chiral COFs through a linker decomposition chiral induction (LDCI) strategy. Three pairs of azine‐linked chiral COFs are successfully synthesized by the condensation reactions of C3‐symmetric 4,4′,4′′‐(1,3,5‐triazine‐2,4,6‐triyl)tribenzaldehyde (Tz) with flexible chiral dihydrazide linkers derived from malic acid, aspartic acid and tartaric acid, respectively. Remarkably, upon complete or partial decomposition from flexible chiral dihydrazides to hydrazine during COF synthesis, the homochirality of these COFs, originating from the single‐handedness conformation of propeller‐like Tz cores, is well preserved. Such a stereoselective chiral memory realized via the LDCI strategy is confirmed by time‐dependent powder X‐ray diffraction (PXRD), Fourier transform infrared (FT‐IR) and diffuse reflectance circular dichroism (DRCD). Moreover, the resultant azine‐linked chiral COFs are used as the active materials to fabricate photodetectors to directly distinguish circularly polarized light (CPL), showing impressive recognition performances on the identification of left‐handed circularly (LHC) and right‐handed circularly (RHC) polarized lights. Notably, the residual undecomposed flexible chiral linkers within the COFs are found to be conducive to improving the polarization discrimination ratio. This work highlights LDCI as a new and effective strategy for constructing homochiral COFs with promising future in chiral optical application.

Application of metabolomics approach to investigate the flavor substance differences between triploid and diploid oysters (Crassostrea angulata)

Oysters, particularly Portuguese oyster (Crassostrea angulata), are highly valued for their nutritional and flavor qualities, making them important in global aquaculture. Triploid oysters have gained attention for maintaining higher meat quality year-round compared to diploids, but there is limited research on how ploidy affects their biochemical and flavor profiles. This study uses a non-targeted metabolomics approach, including gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS), to investigate flavor substance differences between triploid and diploid C. angulata. A total of 13 volatile compounds were identified in diploid oysters, while 28 were found in triploids. Significant upregulation of inosine, guanosine, L-aspartic acid, and taurine in triploids contributes to their enhanced flavor profile. Additionally, triploids showed higher nicotinamide concentrations, while diploids had increased 25-hydroxycholesterol. These findings highlight the advantages of triploid oysters in aquaculture for improved flavor and nutrition, supporting their potential for year-round production.

Comparative Study of Amino Acids in the Life Stages of Silkworm (Bombyx mori L.)

The silkworm (Bombyx mori L.) undergoes distinct life stages, including egg, larva (caterpillar), pupa (chrysalis), and adult moth(imago), each with specific physiological requirements. Amino acids are essential for the growth and development of the silkworm. In this study, we work in paper chromatography to compare the amino acid profiles in different life stages of the silkworm. Samples were collected from eggs, larvae, pupae, and adult moths. Amino acids were extracted and separated on chromatographic paper using a suitable solvent system. The developed chromatograms were analyzed to determine the relative concentrations of specific amino acids. Our results revealed variations in the amino acid composition across the life stages. The amino acids such as glycine, methionine, leucine, and aspartic acid were common in all life stages of silkworms. These differences may be attributed to the physiological changes associated with metamorphosis and reproduction in adult silkworms. Overall, this study provides valuable insights into the dynamic changes in amino acid profiles during the life stages of silkworms, highlighting the importance of amino acids in their growth, development, and silk production. And, this study demonstrates the utility of paper chromatography as a cost-effective and reliable method for analyzing amino acid composition in complex biological samples, providing valuable insights into the nutritional dynamics of the silkworm lifecycle.

Petunidin attenuates vinclozolin instigated testicular toxicity in albino rats via regulating TLR4/MyD88/TRAF6 and Nrf-2/Keap-1 pathway: A pharmacodynamic and molecular simulation approach

Vinclozolin (VZN) is a widely used fungicide which exerts deleterious impacts on various organs including testis. Petunidin (PDN) is a polyphenolic compound that demonstrates a broad range of pharmacological activities. Thirty-two rats were divided into 4 groups including the control, VZN (100 mg/kg), VZN (100 mg/kg) + PDN (4 mg/kg) and PDN (4 mg/kg) treated group. The activities of antioxidant enzymes were assessed by using previously documented protocols. The gene expressions were determined by using qRT-PCR. The levels of hepatic function and apoptotic markers were evaluated by using standard ELISA technique. The histological analysis was carried out as per the standard protocol of histology. It was revealed that VZN disrupted the Nrf-2/Keap-1 pathway. Moreover, the activities of catalase (CAT), glutathione peroxidase (GPx), superoxide dismutase (SOD), heme-oxygenase-1 (HO-1) and glutathione reductase (GSR) were reduced whereas levels of reactive oxygen species (ROS) & malondialdehyde (MDA) were promoted following the VZN intoxication. Furthermore, VZN intoxication reduced total sperm count, viability, motility as well as luteinizing hormone (LH), follicle stimulating hormone (FSH), and plasma testosterone. Besides, administration of VZN decreased the expressions of 3β-Hydroxysteroid dehydrogenase (3β-HSD), steroidogenic acute regulatory protein (StAR) and 17β-Hydroxysteroid dehydrogenase (17β-HSD). Moreover, VZN exposure escalated the expressions of Bcl-2–associated X protein (Bax) and cysteine–aspartic acid protease-3 (Caspase-3) while reducing the expressions of B-cell lymphoma-2 (Bcl-2). Additionally, VZN administration increased the gene expression of toll-like receptor 4 (TLR4), tumor necrosis factor receptor-associated factor 6 (TRAF-6) and myeloid differentiation primary response 88 (MyD88). The levels of interleukin-6 (IL-6), nuclear factor kappa-B (NF-κB), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and the activity of cyclooxygenase-2 (COX-2) were promoted following the VZN administration. Furthermore, VZN intoxication disrupted the normal histology of testicular tissues. However, VZN + PDN treatment ameliorated testicular damage via regulating aforementioned dysregulations owing to its anti-inflammatory, antioxidative as well as anti-apoptotic potentials. Lastly, molecular docking (MD) was performed to assess the effectiveness of PDN as a curative compound by analyzing its binding affinity with the targeted proteins (Keap1, TLR4 and StAR). Our in-silico evaluations confirmed that PDN possesses the potential to interact with binding pockets of these proteins, emphasizing its capability as a curative compound to mitigate VZN-prompted reproductive damage.

Amyloid-like Aggregation Propensities of Metabolites- Homogentisic Acid, N-Acetyl Aspartic Acid and Isovaleric Acid.

The transformation of metabolites into amyloidogenic aggregates represent an intriguing dimension in the pathophysiology of metabolic disorders, including alkaptonuria, canavan disease, and isovaleric acidemia. Central to this phenomenon are the metabolites homogentisic acid (HA), N-acetyl aspartic acid (NAA), and isovaleric acid (IVA), which we found, weave an intricate network of self-assembled structures. Leveraging an array of microscopy techniques, we traced the morphological behavior of these assemblies that exhibit concentration and time-dependent morphological transitions from isolated globules to clustered aggregates. MD simulation studies suggest significant role of hydrogen bonding interactions in the aggregation process. While displaying strong amyloidogenic propensity in solution, these aged aggregates were significantly cytotoxic to mouse neural N2a cell lines. In vivo effect in Caenorhabditis elegans (C. elegans) nematode further validated cytotoxicity of aggregates. Our findings provide fresh insights to amyloidogenic nature of HA, NAA, and IVA aggregates and their possible role in associated metabolic disorders such as alkaptonuria, canavan disease and isovaleric acidemia.

Contrasting the Effects of Aspartic Acid and Glycine in Free Amino Acid and Peptide Forms on the Growth Rate, Morphology, Composition, and Structure of Synthetic Aragonites

Contrasting the Effects of Aspartic Acid and Glycine in Free Amino Acid and Peptide Forms on the Growth Rate, Morphology, Composition, and Structure of Synthetic Aragonites

Research on the properties of crystalline admixtures: Self-healing healing materials for concrete from multiple perspectives

The research on the self-healing of concrete promoted by crystal admixtures provides a new direction for the green development of construction industry. In this study, three crystal admixtures, i.e., l-aspartic acid (LAA), Na2SiO3 and nano-silica (NS), were screened from the experimental results in terms of promotion of hydration, fast reaction, and effective filling. The three components were mixed as concrete self-healing agents, and the experimental ratio was designed by response surface method. Finally, the optimal ratio was selected according to the visual healing effect. The durability of mortar with the optimum ratio of self-healing agent was studied by the experiment of resistance to chloride ion and sulfate attack; its preliminary heat release was studied by hydration microcalorimeter. The microscopic analysis was carried out by X-ray powder diffraction (XRD), thermogravimetric analysis (TG), scanning electron microscopy (SEM), Brunauer-Emmett-Teller (BET) test and mercury injection method (MIP). The experimental results showed that the best healing effect was achieved when the dosage of LAA, Na2SiO3 and NS reached 0.5 %, 0.4 % and 0.15 % of the cement mass. In the process of healing, the early healing products are mainly calcium carbonate, and the late healing products are mainly ettringite and calcium silicate hydrate (CSH), and the healing agent effectively reduces the porosity and improves the pore structure, making the concrete matrix more dense.

Measurement of circular intensity differential scattering (CIDS) from single optically trapped biological particles

The circular intensity differential scattering (CIDS), which is the normalized Mueller matrix element -S14/S11, has been measured from single biological particles as a function of scattering angle. CIDS is valuable for its potential in detecting chiral particles that may include the helical structures of DNA or RNA molecules in biological samples, and as such is a potential method for detecting biological particles. Optical trapping is employed to levitate single particles within a custom-designed elliptical reflector for CIDS measurements. The advantage of optical levitation in light-scattering measurements is that single particles can be suspended in air with sufficient working distance to prevent interference from the suspending apparatus. To measure the phase function, the reflector is used to collect the angle-dependent scattering signals. We demonstrated that we can obtain two-dimensional angular optical scattering (TAOS) patterns that cover a wide angular range from single levitated particles. These TAOS patterns are generated using 532 nm illumination of left-handed and right-handed circular polarizations and recorded from trapped single particles (silica, English Oak, Ragweed, Mulberry, Glycine, and l-Aspartic acid).

Comprehensive studies of the serine carboxypeptidase-like (SCPL) gene family in Carya cathayensis revealed the roles of SCPL4 in epigallocatechin-3-gallate (EGCG) synthesis and drought tolerance

Hickory (Carya cathayensis) nuts are rich in epigallocatechin-3-gallate (EGCG) with multiple health functions. EGCG also regulates plant growth, development and stress responses. However, research on the synthesis mechanism of EGCG and its function in hickory is currently limited. Herein, 44 serine carboxypeptidase-like (SCPL) members were identified from the hickory genome and classified into three major categories: SCPL-I, SCPL-II, and SCPL-III. In the CcSCPLs-IA branch, CcSCPL3/4/5/8/9/11/13 showed differential expression patterns in various tissues, especially with relatively high expression levels in plant roots, female flowers and seed coat. These proteins have a catalytic triad composed of serine (Ser), aspartic acid (Asp) and histidine (His). Ser-His in the triad and arginine (Arg) mediated the docking of CcSCPL3/4/5/11 with 1-O-galloyl-β-d-glucose (βG) and epigallocatechin (EGC), whereas the Asp of the triad did not. CcSCPL4 was further confirmed to promote the synthesis of EGCG in tobacco leaves. CcSCPL4 may function as monomer and be mainly localized within cellular structures outside the nucleus. Notably, the expression level of CcSCPL4 significantly changed after drought, cold, and salt stress, with the highest expression level under drought stress. Meanwhile CcSCPL4 over-expression could enhance the drought resistance of Saccharomyces cerevisiae and Arabidopsis. This study elucidates key enzymes for EGCG synthesis and their role in drought resistance, providing insights into the EGCG synthesis pathway and molecular breeding of hickory in future.

Effect of Extraction Processes on the Physicochemical and Functional Properties of Protein-Rich Extracts from the Red Seaweed Pyropia Seriata (Nori)

This study examined the physicochemical and techno-functional properties of protein extracts from the red seaweed (Pyropia seriata). The extraction processes involved variations of some of the following steps: alkaline treatment, milling, freeze drying, neutralisation, heating, and concentration. The extracts had total amino acid contents ranging between 40–55% of the powders with essential amino acids comprising 25–30% of the powders. The extracts were particularly abundant in aspartic acid and limited in the essential sulphur amino acids. The molecular weight (MW) distribution of the proteins within the extracts ranged from <10 to 50 kDa. All the extracts were soluble, with a solubility of 65–100% depending on the final pH of the solution. The extracts had significant foaming-producing and emulsifying potential. The foaming stability ranged between 75–91% for solutions between pH 7 to 11 and less than 75% for solutions with pH of 5 or 3. Alkaline treatment significantly improved foaming capacity (120-160%). A similar effect was also observed for emulsifying activity (0.29–0.51) and emulsifying stability (15–40%). Milling enhanced the emulsifying potential and decreased the mg of amino acids per g powder. Neutralisation with hydrochloric acid instead of citric acid and subsequent heating and concentration showed minimal effect on the properties of the extracts and decreased the mg amino acids per g powder. This research highlights the potential of Pyropia seriata as a source of soluble alternative protein for food applications.

Antioxidant hydrogel from poly(aspartic acid) and carboxymethylcellulose with quercetin loading as burn wound dressing

Susceptibility to infection and excessive accumulation of reactive oxygen species (ROS) are the greatest obstacles for burn wound healing. In this research, the 5-aminosalicylic acid (ASA) grafted poly(aspartic hydrazide) (PASH) was synthesized by successive ploysuccinimide (PSI) ring opening reaction and reacted with oxidized carboxymethyl cellulose (DCMC) to fabricate biodegradable hydrogel through Schiff-base cross-linking. Moreover, the hydrogel was loaded with quercetin (QT) to enhance its anti-inflammatory performance. The ASA moiety endowed the hydrogel with the free radical scavenging ability and mussel inspired tissue adhesion to maintain the healing bioenvironment of the wound. The loading of QT gave the hydrogel more phenolic hydroxy group and further enhanced the antioxidant capacity of the hydrogel. The in vitro experiment revealed the grafted ASA moiety and the loaded QT greatly enhanced the ROS elimination property and antibacterial property. Moreover, the QT loaded hydrogel accelerated the burn wound repairing rate in the in vivo mice model. Based on above result, the PASH/DCMC could act as a new platform for QT loading to promote the burn wound repairing.

Preliminary study on the potential impact of probiotic combination therapy on Helicobacter pylori infection in children using 16S gene sequencing and untargeted metabolomics approach

ObjectiveThe purpose of this study was to explore the potential mechanism of Helicobacter pylori (Hp) eradication by probiotic therapy through 16S rRNA gene sequencing technology and untargeted metabolomics.MethodsTwenty four Hp-infected children were recruited from the Shanxi Bethune Hospital, and 24 healthy children were recruited as a blank control group. Group A: fecal samples from 24 healthy children. Group B: fecal samples of 24 children with Hp infection. Group B1 (n = 15): fecal samples of group B treated with probiotic therapy for 2 weeks. Group B2 (n = 19): fecal samples of group B treated with probiotic therapy for 4 weeks. The above fecal samples were analyzed by 16S rRNA gene sequencing technology and untargeted metabolomics.ResultsThere was no significant difference in alpha diversity and beta diversity among the four groups, but many bacteria with statistical difference were found in each group at the bacterial genus level and phylum level. LEfSe results showed that in group B, Porphyromonadaceae, Shigella and other microorganisms related to intestinal microecological dysbiosis were enriched. And in group B2, abundant characteristic microorganisms were found, namely Bacillales and Prevotella. KEGG metabolic pathway enrichment analysis showed that groups B1 and B2 were involved in 10 metabolic pathways potentially related to probiotic treatment: purine metabolism, nitrogen metabolism, arginine biosynthesis, alanine, aspartic acid and glutamate metabolism, glyoxylic acid and dicarboxylic acid metabolism, unsaturated fatty acid biosynthesis, fatty acid extension, fatty acid degradation, pyrimidine metabolism, fatty acid biosynthesis.ConclusionProbiotic therapy can inhibit Hp to some extent and can relieve gastrointestinal symptoms, making it a preferred therapy for children with Hp infection and functional abdominal pain. Hp infection can reduce the diversity of intestinal microbes, resulting in the disturbance of intestinal microbiota and changes in the relative abundance of microbiota in children, while probiotic therapy can restore the diversity of intestinal microbes and intestinal microecological balance.

Uncovering the taste features: Applying machine learning and molecular docking approaches to predict umami taste intensity of peptides

The umami taste, often described as the fifth basic taste, plays a pivotal role in the culinary world, significantly contributing to the overall flavor profile and consumer satisfaction of food products. Precise prediction and enhancement of umami taste intensity using the identification of umami peptides can lead to groundbreaking advancements in the food industry. This study presents a novel approach to combine machine learning and molecular docking techniques. The machine learning algorithm is based on a cascade algorithm combining CatBoost and BERT models to predict the umami taste intensity of peptides accurately. Our research reveals that combinations of specific amino acids, such as aspartic acid with alanine or glycine and lysine with glycine or histidine, have been identified to enhance the umami taste in foods. The model is available as a user-friendly web server at https://taste.infochemistry.ru. This study contributes to the scientific understanding of taste perception and provides a valuable tool for the food industry to innovate and improve product quality by optimizing umami taste profiles.

Evaluation of Selected Plant Phenolics via Beta-Secretase-1 Inhibition, Molecular Docking, and Gene Expression Related to Alzheimer’s Disease

Background: The goal of the current study was to investigate the inhibitory activity of six phenolic compounds, i.e., rosmarinic acid, gallic acid, oleuropein, epigallocatechin gallate (EGCG), 3-hydroxytyrosol, and quercetin, against β-site amyloid precursor protein cleaving enzyme-1 (BACE1), also known as β-secretase or memapsin 2, which is implicated in the pathogenesis of Alzheimer’s disease (AD). Methods and Results: The inhibitory potential against BACE1, molecular docking simulations, as well as neurotoxicity and the effect on the AD-related gene expression of the selected phenolics were tested. BACE1 inhibitory activity was carried out using the ELISA microplate assay via fluorescence resonance energy transfer (FRET) technology. Molecular docking experiments were performed in the human BACE1 active site (PDB code: 2WJO). Neurotoxicity of the compounds was carried out in SH-SY5Y, a human neuroblastoma cell line, by the Alamar Blue method. A gene expression analysis of the compounds on fourteen genes linked to AD was conducted using the real-time polymerase chain reaction (RT-PCR) method. Rosmarinic acid, EGCG, oleuropein, and quercetin (also used as the reference) were able to inhibit BACE1 with their respective IC50 values 4.06 ± 0.68, 1.62 ± 0.12, 9.87 ± 1.01, and 3.16 ± 0.30 mM. The inhibitory compounds were observed to occupy the non-catalytic site of the BACE1. However, hydrogen bonds were found to be present between rosmarinic acid and EGCG and aspartic amino acid D228 in the catalytic site. Oleuropein and quercetin effectively suppressed the expression of PSEN, APOE, and CLU, which are recognized to be linked to the pathogenesis of AD. Conclusions: The outcomes of the work bring quercetin, EGCG, and rosmarinic acid to the forefront as promising BACE1 inhibitors.

Circulating amino acid signature features urea cycle alterations associated with coronary artery disease

Coronary artery disease (CAD) remains a leading cause of death worldwide and imposes a substantial socioeconomic burden on healthcare. Improving risk stratification in clinical practice could help to combat this burden. As amino acids are biologically active metabolites whose involvement in CAD remains largely unknown, this study investigated associations between circulating amino acid levels and CAD phenotypes. A high-coverage quantitative liquid chromatography-mass spectrometry approach was applied to acquire the serum amino acids profile of age- and sex-coarsened-matched patients with CAD (n = 46, 66.9 years, 74.7% male) and healthy individuals (n = 120, 67.4 years, 74.7% male) from the COmPLETE study. Multiple linear regressions were performed to investigate associations between amino acid levels and (a) the health status (CAD vs. healthy), (b) the number of affected coronary arteries, or (c) the left ventricular ejection fraction. Regressions were adjusted for age, sex, daily physical activity, sampling, and fasting time. Urea cycle amino acids (ornithine, citrulline, homocitrulline, aspartate, and arginine) were significantly and negatively associated with CAD, the number of affected coronary arteries, and the left ventricular ejection fraction. Lysine, histidine, and the glutamine/glutamate ratio were also significantly and negatively associated with the CAD phenotypes. Overall, patients with CAD displayed lower levels of urea cycle amino acids, highlighting a potential role for urea cycle amino acid profiling in cardiovascular risk stratification.Trial registrationThe study was registered on https://www.clinicaltrials.gov (NCT03986892) on June 5, 2019.

Cathepsin B- and L-like Protease Activities Are Induced During Developmental Barley Leaf Senescence

Leaf senescence is a developmental process allowing nutrient remobilization to sink organs. Previously cysteine proteases have been found to be highly expressed during leaf senescence in different plant species. Using biochemical and immunoblotting approaches, we characterized developmental senescence of barley (Hordeum vulgare L. var. ‘GemCraft’) leaves collected from 0 to 6 weeks after the onset of flowering. A decrease in total protein and ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) large subunits occurred in parallel with an increase in proteolytic activity measured using the fluorogenic substrates Z-RR-AMC, Z-FR-AMC, and casein labeled with fluorescein isothiocyanate (casein-FITC). Aminopeptidase activity detected with R-AMC peaked at week 3 and then decreased, reaching a low level by week 6. Maximal proteolytic activity with Z-FR-AMC and Z-RR-AMC was detected from pH 4.0 to pH 5.5 and pH 6.5 to pH 7.4, respectively, while two pH optima (pH 3.6 to pH 4.5 and pH 6.5 to pH 7.4) were found for casein-FITC. Compound E-64, an irreversible cysteine protease inhibitor, and CAA0225, a selective cathepsin L inhibitor, effectively inhibited proteolytic activity with IC50 values in the nanomolar range. CA-074, a selective cathepsin B inhibitor, was less potent under the same experimental conditions, with IC50 in the micromolar range. Inhibition by leupeptin and phenylmethylsulfonyl fluoride (PMSF) was weak, and pepstatin A, an inhibitor of aspartic acid proteases, had no effect at the concentrations studied (up to 0.2 mM). Maximal proteolytic activity with the aminopeptidase substrate R-AMC was detected from pH 7.0 to pH 8.0. The pH profile of DCG-04 (a biotinylated activity probe derived from E-64) binding corresponded to that found with Z-FR-AMC, suggesting that the major active proteases are related to cathepsins B and L. Moreover, immunoblotting detected increased levels of barley SAG12 orthologs and aleurain, confirming a possible role of these enzymes in senescing leaves.

Leishmania donovani adenylosuccinate synthetase requires IMP for dimerization and organization of the active site.

Adenylosuccinate synthetase (AdSS), which catalyses the GTP-dependent conversion of inosine monophosphate (IMP) and aspartic acid to succinyl-AMP, plays a major role in purine biosynthesis. In some bacterial AdSS, it is implicated that IMP binding is important to organize the active site, but in certain plant AdSS, GTP performs this role. Here, we report that in Leishmania donovani AdSS, IMP binding favoured dimerization, induced greater conformational change and improved the protein stability more than GTP binding. IMP binding, which resulted in a network of hydrogen bonds, stabilized the conformation of active site loops and brought the switch loop to a closed conformation, which then facilitated GTP binding. Our results provide a basis for designing better inhibitors of leishmanial AdSS.