Alanine Synthesis Research Articles

Origin and Roles of Alanine and Glutamine in Gluconeogenesis in the Liver, Kidneys, and Small Intestine under Physiological and Pathological Conditions.

Alanine and glutamine are the principal glucogenic amino acids. Most originate from muscles, where branched-chain amino acids (valine, leucine, and isoleucine) are nitrogen donors and, under exceptional circumstances, a source of carbons for glutamate synthesis. Glutamate is a nitrogen source for alanine synthesis from pyruvate and a substrate for glutamine synthesis by glutamine synthetase. The following differences between alanine and glutamine, which can play a role in their use in gluconeogenesis, are shown: (i) glutamine appearance in circulation is higher than that of alanine; (ii) the conversion to oxaloacetate, the starting substance for glucose synthesis, is an ATP-consuming reaction for alanine, which is energetically beneficial for glutamine; (iii) most alanine carbons, but not glutamine carbons, originate from glucose; and (iv) glutamine acts a substrate for gluconeogenesis in the liver, kidneys, and intestine, whereas alanine does so only in the liver. Alanine plays a significant role during early starvation, exposure to high-fat and high-protein diets, and diabetes. Glutamine plays a dominant role in gluconeogenesis in prolonged starvation, acidosis, liver cirrhosis, and severe illnesses like sepsis and acts as a substrate for alanine synthesis in the small intestine. Interactions among muscles and the liver, kidneys, and intestine ensuring optimal alanine and glutamine supply for gluconeogenesis are suggested.

Mechanisms of interstellar synthesis of glycine, alanine, and serine from aminonitriles, OH, and H2O

AbstractA new potential energy surface for forming glycine in the gas phase, starting from association of aminoacetonitrile (NH2CH2CN) with OH followed by subsequent hydrolysis, was determined using CBS‐QB3 calculation. The overall activation energy was 90 kJ mol−1 or 0 without or with catalytic H2O, respectively. Alanine or serine was formed from 2‐aminopropionitrile (CH3CH(NH2)CN) or 2‐amino‐3‐hydrxypropionitrile (HOCH2CH(NH2)CN) instead of aminoacetonitrile with the overall activation energies of 89 or 49 kJ mol−1, respectively. When an additional H2O was involved in each reaction as a catalyst, barrierless reaction pathways were obtained. These results suggest that it is possible for investigated reactions to occur in interstellar ices along the proposed pathways, taking kinetic aspects into account.

The Geochemical Potential for Metabolic Processes on the Sub-Neptune Exoplanet K2-18b

Quantifying disequilibria is important to understand whether an environment could be habitable. It has been proposed that the exoplanet K2-18b has a hydrogen-rich atmosphere and a water ocean, making it a “hycean world.” The James Webb Space Telescope recently made measurements of methane, CO2, and possibly dimethyl sulfide (DMS) in the atmosphere of this planet. The initial interpretation of these data is that they may support the occurrence of hycean conditions. Here I attempt to take a next step in exploring the prospects for habitability. I use constraints on the abundances of atmospheric gases to calculate how much chemical disequilibrium there could be, assuming that K2-18b is a hycean world. I find that the presence of oxidized carbon species coexisting with abundant H2 (1–1000 bars) at cool to warm (25°C–120°C) conditions creates a strong thermodynamic drive for methanogenesis. More than ∼75 kJ (mol C)−1 of free energy can be released from CO2 hydrogenation. Partially oxidized carbon compounds such as DMS (if present) also have the potential to provide metabolic energy, albeit in smaller quantities. Because of the thermodynamic instability of CO2 under hycean conditions, other reductive reactions of CO2 are likely to be favored, including the synthesis of amino acids. Glycine and alanine synthesis can be energy releasing or at least much less costly on K2–18b than in Earth’s ocean, even when NH3 is scarce but not totally absent. These first bioenergetic calculations for a proposed ocean-bearing exoplanet lay new groundwork for assessing exoplanetary habitability.

A metabolomics perspective on 2 years of high-intensity training in horses

The plasma metabolomic profile of elite harness horses subjected to different training programmes was explored. All horses had the same training programme from 1.5 until 2 years of age and then high-intensity training was introduced, with horses divided into high and low training groups. Morning blood samples were collected at 1.5, 2, 2.5 and 3.5 years of age. The plasma was analysed using targeted absolute quantitative analysis and a combination of tandem mass spectrometry, flow-injection analysis and liquid chromatography. Differences between the two training groups were observed at 2 years of age, when 161 metabolites and sums and ratios were lower (e.g. ceramide and several triglycerides) and 51 were higher (e.g. aconitic acid, anserine, sum of PUFA cholesteryl esters and solely ketogenic AAs) in High compared with low horses. The metabolites aconitic acid, anserine, leucine, HArg synthesis and sum of solely ketogenic AAs increased over time, while beta alanine synthesis, ceramides and indole decreased. Therefore high-intensity training promoted adaptations linked to aerobic energy production and amino acid metabolism, and potentially also affected pH-buffering and vascular and insulin responses.

Integrated analysis of physiological, transcriptome, and metabolome analyses of the gills in Solenaia oleivora under ammonia exposure

Ammonia is a common toxicant in aquatic systems and one of the key factors affecting aquaculture. However, data on mollusks’ toxic response and coping mechanisms to ammonia nitrogen, especially freshwater mollusks, are still lacking. In this study, we evaluated the tolerance of a freshwater mollusk Solenaia oleivora to ammonia and investigated its coping mechanisms by combining physiological, metabolic, and transcriptomic analyses in the gills. The acute toxicity test revealed that the LC50–96 h (temperature-20 ℃, pH-7.4) of ammonia in S. oleivora was 63.29 mg/L. The physiological and TUNEL results showed that although 10 mg/L ammonia exposure increased the activities of antioxidant, immune and ammonia detoxification-related enzymes, it still caused oxidative damage and cell apoptosis of gill tissues. A total of 97 differential metabolites (DMs) and 3431 differential expressed genes (DEGs) were identified after ammonia stress. Among them, most DMs and DEGs were involved in immune response, antioxidant, cell apoptosis, carbohydrate metabolism, amino acid metabolism, and lipid metabolism. The enhancement of glycolysis and lipid metabolisms may provide energy for immune response and ammonia detoxification. In addition, glutamine synthesis, alanine synthesis and urea cycle were involved in ammonia nitrogen detoxification in the gill tissue of S. oleivora. Our results indicate that ammonia leads to individual death in S. oleivora, as wells as oxidative damage, cell apoptosis, immune response, and metabolic changes of gill tissues. The findings will provide valuable information to assess the potential ecological risk of environmental ammonia to freshwater mollusks and theoretical guidance for the healthy aquaculture of S. oleivora.

ERCC6L2 Disease and Shwachman-Diamond Syndrome Exhibit Alterations in Metabolomes in Patient Fibroblasts

Introduction Metabolomics is an emerging tool to study metabolic alterations in cancers. It has also the potential to identify clinically relevant biomarkers for diagnosis, risk appraisal, and targeted drug development. In hematology, metabolic reprogramming is essential for normal regulation of hematopoiesis. Association of discrete metabolic alterations with explicit somatic mutations have also been reported in hematological malignancies. We have previously observed glutaminase (GLS) overexpression and improved oxygen consumption rate with excess glutamine in low glucose conditions in patient-derived fibroblasts in ERCC6L2 disease (unpublished). This prompted us to study metabolomic alterations in ERCC6L2 disease and Shwachman-Diamond syndrome (SDS) induced strictly by the germline ERCC6L2 and SBDS mutations. Both inherited bone marrow failure (BMF) syndromes carry considerable tendency of somatic TP53 mutagenesis predisposing patients to a high risk myeloid malignancy. Our aim here was to identify potential drug targets by targeted metabolomics. Methods We examined patient-derived skin fibroblasts (ERCC6L2 disease, n=4; SDS, n=4) and healthy controls (n=4) and performed targeted metabolomics with U-13C-D-glucose and U-13C-L-glutamine labeling. Fibroblasts were seeded on six-well plates and on the following day exposed to 13C-glucose (25mM) or 13C-glutamine (2mM) label containing medium for 30 or 120 minutes. Next, metabolites were extracted and quantified with liquid chromatography mass spectrometer. The results were normalized to total metabolite intensity per sample. Results In 13C-glucose labeled samples, we detected statistically significantly lower intensity of pyruvate M+3 in both ERCC6L2 and SDS fibroblasts compared to control at 30 minutes, which persisted at 120 minutes in SDS cells. In the TCA cycle, malate M+3 labeling indicates pyruvate carboxylase (PC) activity and M+2 labeling pyruvate dehydrogenase activity (PDH) (Figure 1). We found a significantly higher portion of malate M+3 than M+2 in SDS cells at all time points, suggesting lower PDH and higher PC activity in SDS cells. Additionally, we detected higher levels of alanine M+3 in SDS samples. As pyruvate is a source for alanine synthesis, we suspect an alteration in pyruvate metabolism; potentially to overcome challenges in pyruvate oxidation due to either decrease in the respiratory chain function or PDH activity. In 13C-glutamine labeled samples, we detected statistically significantly higher intensity of glutamine M+5 and M+4 in ERCC6L2 samples at 30 minutes compared to control and SDS cells. Glutamine is first converted to glutamate by glutaminase (GLS), after which it enters the TCA cycle as ketoglutarate (KG) (Figure 1). In addition to glutamine, we detected an increase, although not statistically significant, in glutamate M+5 and M+4, as well as KG M+5, in ERCC6L2 cells at 30 minutes. A similar trend was observed in the subsequent TCA cycle intermediates succinic acid M+4, fumarate M+4, and malate M+4. At 120 minutes, there was no difference in metabolite intensities between the groups, suggesting a potentially faster glutamine uptake by ERCC6L2 cells. Conclusions Our analysis using non-hematopoietic tissue provides understanding of metabolic weak links not confounded by somatic events occurring early in the disease course in ERCC6L2 and SDS patients. Our findings imply increased glutamine utilization in ERCC6L2 cells and altered pyruvate metabolism in SDS cells. Therefore, these metabolic pathways are attractive drug targets (summarized in Figure 1). Further studies are needed to determine their potential in supporting the compromised hematopoiesis to reduce the stress for somatic mutagenesis.

Integrated Tandem Electrochemical‐chemical‐electrochemical Coupling of Biomass and Nitrate to Sustainable Alanine

AbstractAlanine is widely employed for synthesizing polymers, pharmaceuticals, and agrochemicals. Electrocatalytic coupling of biomass molecules and waste nitrate is attractive for the nitrate removal and alanine production under ambient conditions. However, the reaction efficiency is relatively low due to the activation of the stable substrates, and the coupling of two reactive intermediates remains challenging. Herein, we realize the integrated tandem electrochemical‐chemical‐electochemical synthesis of alanine from the biomass‐derived pyruvic acid (PA) and waste nitrate (NO3−) catalyzed by PdCu nano‐bead‐wires (PdCu NBWs). The overall reaction pathway is demonstrated as a multiple‐step catalytic cascade process via coupling the reactive intermediates NH2OH and PA on the catalyst surface. Interestingly, in this integrated tandem electrochemical‐chemical‐electrochemical catalytic cascade process, Cu facilitates the electrochemical reduction of nitrate to NH2OH intermediates, which chemically couple with PA to form the pyruvic oxime, and Pd promotes the electrochemical reduction of pyruvic oxime to the desirable alanine. This work provides a green strategy to convert waste NO3− to wealth and enriches the substrate scope of renewable biomass feedstocks to produce high‐value amino acids.

Integrated Tandem Electrochemical-chemical-electrochemical Coupling of Biomass and Nitrate to Sustainable Alanine.

Alanine is widely employed for synthesizing polymers, pharmaceuticals, and agrochemicals. Electrocatalytic coupling of biomass molecules and waste nitrate is attractive for the nitrate removal and alanine production under ambient conditions. However, the reaction efficiency is relatively low due to the activation of the stable substrates, and the coupling of two reactive intermediates remains challenging. Herein, we realize the integrated tandem electrochemical-chemical-electochemical synthesis of alanine from the biomass-derived pyruvic acid (PA) and waste nitrate (NO3-) catalyzed by PdCu nano-bead-wires (PdCu NBWs). The overall reaction pathway is demonstrated as a multiple-step catalytic cascade process via coupling the reactive intermediates NH2OH and PA on the catalyst surface. Interestingly, in this integrated tandem electrochemical-chemical-electrochemical catalytic cascade process, Cu facilitates the electrochemical reduction of nitrate to NH2OH intermediates, which chemically couple with PA to form the pyruvic oxime, and Pd promotes the electrochemical reduction of pyruvic oxime to the desirable alanine. This work provides a green strategy to convert waste NO3- to wealth and enriches the substrate scope of renewable biomass feedstocks to produce high-value amino acids.

Synthesis, DFT study, molecular docking and larvicidal activity of Chitin inhibitor alanine derived compounds

In order to obtain an effective larvicidal against A. aegypti the vectors for Dengue, Zika and Chikungunya virus, Sodium 2-{(E)-[(2-hydroxy-3-methoxyphenyl) methylidene] amino} propanoate (NaLH) ligand and its Cu(II) complexes derived from alanine amino acid with and without coligand (C1 &C2) have been synthesized. All three molecules were characterized by physicochemical techniques including FTIR, UV–Vis, proton NMR and Mass spectrum. The geometry optimization and frontier orbitals HOMO, LUMO and band gaps were calculated by using B3LYP functional and SVP basis sets for all three compounds (NaLH, complex C1 and C2). The values of HOMO/LUMO energies observed were −4.2/−1.7 eV for ligand, −5.2–3.5 eV for C1 and −5.0/−3.7 eV for C2 and corresponding values of energy gaps were 2.5 eV, 1.7 eV and 1.3 eV for NaLH, C1 and C2 respectively which increases in the following order NaLH > C1 > C2. Complex1 has been arranged in slightly distorted octahedral geometry while complex2 acquired a square pyramidal geometry. Molecular docking study of the reported larvicidal against insect acetylcholistenerase (PDB id: 6ARY) and Chitin acetylase (PDB id 5ZNT) revealed significant free energy of binding in the active site of target molecules with binding energy of −6.14 & −10.20Kcal/mol and −6.79 & −11.75Kcal/mol for C1 &C2 respectively. In addition, Larvicidal activity of complex 2 was observed with 100% mortality rate in just 2 hrs time using 1.0% concentration of larvicide. The LC50 value is 0.0015 µg/mL while LC90 value is 0.0018 µg/mL for C2. The complexes were observed to be lethal against Chitinous body layer of larvae of Aedes aegypti, the vectors for Dengue, Zika and Chikungunya virus.

Lignin-derived carbon nanosheets boost electrochemical reductive amination of pyruvate to alanine

Efficient and sustainable amino acid synthesis is essential for industrial applications. Electrocatalytic reductive amination has emerged as a promising method, but challenges such as undesired side reactions and low efficiency persist. Herein, we demonstrated a lignin-derived catalyst for alanine synthesis. Carbon nanosheets (CNSs) were synthesized from lignin via a template-assisted method and doped with nitrogen and sulfur to boost reductive amination and suppress side reactions. The resulting N,S-co-doped carbon nanosheets (NS-CNSs) exhibited outstanding electrochemical performance. It achieved a maximum alanine Faradaic efficiency of 79.5%, and a yield exceeding 1,199μmol h-1 cm-2 on NS-CNS, with a selectivity above 99.9%. NS-CNS showed excellent durability during long-term electrolysis. Kinetic studies including control experiments and theoretical calculations provided further insights into the reaction pathway. Moreover, NS-CNS catalysts demonstrated potential in upgrading real-world polylactic acid plastic waste, yielding value-added alanine with a selectivity over 75%.

Rational metabolic engineering of Corynebacterium glutamicum for efficient synthesis of L-glutamate

L-glutamic acid is the world's largest bulk amino acid product that is widely used in the food, pharmaceutical and chemical industries. Using Corynebacterium glutamicum G01 as the starting strain, the fermentation by-product alanine content was firstly reduced by knocking out the gene encoding alanine aminotransferase (alaT), a major by-product related to alanine synthesis. Secondly, since the α-ketoglutarate node carbon flow plays an important role in glutamate synthesis, the ribosome-binding site (RBS) sequence optimization was used to reduce the activity of α-ketoglutarate dehydrogenase and enhance the glutamate anabolic flow. The endogenous conversion of α-ketoglutarate to glutamate was also enhanced by screening different glutamate dehydrogenase. Subsequently, the glutamate transporter was rationally desgined to improve the glutamate efflux capacity. Finally, the fermentation conditions of the strain constructed using the above strategy were optimized in 5 L fermenters by a gradient temperature increase combined with a batch replenishment strategy. The glutamic acid production reached (135.33±4.68) g/L, which was 41.2% higher than that of the original strain (96.53±2.32) g/L. The yield was 55.8%, which was 11.6% higher than that of the original strain (44.2%). The combined strategy improved the titer and the yield of glutamic acid, which provides a reference for the metabolic modification of glutamic acid producing strains.

Genome Analysis and Optimization of Caproic Acid Production of Clostridium butyricum GD1-1 Isolated from the Pit Mud of Nongxiangxing Baijiu.

Caproic acid is a precursor substance for the synthesis of ethyl caproate, the main flavor substance of nongxiangxing baijiu liquor. In this study, Clostridium butyricum GD1-1, a strain with high caproic acid concentration (3.86 g/l), was isolated from the storage pit mud of nongxiangxing baijiu for sequencing and analysis. The strain's genome was 3,840,048 bp in length with 4,050 open reading frames. In addition, virulence factor annotation analysis showed C. butyricum GD1-1 to be safe at the genetic level. However, the annotation results using the Kyoto Encyclopedia of Genes and Genomes Automatic Annotation Server predicted a deficiency in the strain's synthesis of alanine, methionine, and biotin. These results were confirmed by essential nutrient factor validation experiments. Furthermore, the optimized medium conditions for caproic acid concentration by strain GD1-1 were (g/l): glucose 30, NaCl 5, yeast extract 10, peptone 10, beef paste 10, sodium acetate 11, L-cysteine 0.6, biotin 0.004, starch 2, and 2.0% ethanol. The optimized fermentation conditions for caproic acid production by C. butyricum GD1-1 on a single-factor basis were: 5% inoculum volume, 35°C, pH 7, and 90% loading volume. Under optimal conditions, the caproic acid concentration of strain GD1-1 reached 5.42 g/l, which was 1.40 times higher than the initial concentration. C. butyricum GD1-1 could be further used in caproic acid production, NXXB pit mud strengthening and maintenance, and artificial pit mud preparation.

Gut microbiome signatures of extreme environment adaption in Tibetan pig

Tibetan pigs (TPs) can adapt to the extreme environments in the Tibetan plateau implicated by their self-genome signals, but little is known about roles of the gut microbiota in the host adaption. Here, we reconstructed 8210 metagenome-assembled genomes from TPs (n = 65) living in high-altitude and low-altitude captive pigs (87 from China—CPs and 200 from Europe—EPs) that were clustered into 1050 species-level genome bins (SGBs) at the threshold of 95% average nucleotide identity. 73.47% of SGBs represented new species. The gut microbial community structure analysis based on 1,048 SGBs showed that TPs was significantly different from low-altitude captive pigs. TP-associated SGBs enabled to digest multiple complex polysaccharides, including cellulose, hemicellulose, chitin and pectin. Especially, we found TPs showed the most common enrichment of phyla Fibrobacterota and Elusimicrobia, which were involved in the productions of short- and medium-chain fatty acids (acetic acid, butanoate and propanoate; octanomic, decanoic and dodecanoic acids), as well as in the biosynthesis of lactate, 20 essential amino acids, multiple B vitamins (B1, B2, B3, B5, B7 and B9) and cofactors. Unexpectedly, Fibrobacterota solely showed powerful metabolic capacity, including the synthesis of acetic acid, alanine, histidine, arginine, tryptophan, serine, threonine, valine, B2, B5, B9, heme and tetrahydrofolate. These metabolites might contribute to host adaptation to high-altitude, such as energy harvesting and resistance against hypoxia and ultraviolet radiation. This study provides insights into understanding the role of gut microbiome played in mammalian high-altitude adaptation and discovers some potential microbes as probiotics for improving animal health.

Enantiodetermining processes in the synthesis of alanine, serine, and isovaline.

In this study, quantum chemical calculations were used to explore the synthesis of three chiral α-amino acids, specifically alanine, serine, and isovaline, from reactants found in interstellar space. Our focus is on the crucial step in the synthesis pathway that determines the chirality of the amino acids. The results indicate that in the case of alanine, the determination of enantiomer is primarily influenced by the direction of the collision of molecules or functional groups, which leads to the formation of a chirality center in a crucial intermediate. However, contrary to chemical expectations, the enantiodetermining/enantioselection step for serine and isovaline synthesis occurs prior to the creation of a chirality center.

Jian-Ti-Kang-Yi decoction alleviates poly(I:C)-induced pneumonia by inhibiting inflammatory response, reducing oxidative stress, and modulating host metabolism.

Jian-Ti-Kang-Yi decoction (JTKY) is widely used in the treatment of COVID-19. However, the protective mechanisms of JTKY against pneumonia remain unknown. In this study, polyinosinic-polycytidylic acid (poly(I:C)), a mimic of viral dsRNA, was used to induce pneumonia in mice; the therapeutic effects of JTKY on poly(I:C)-induced pneumonia model mice were evaluated. In addition, the anti-inflammatory and anti-oxidative potentials of JTKY were also investigated. Lastly, the metabolic regulatory effects of JTKY in poly(I:C)-induced pneumonia model mice were studied using untargeted metabolomics. Our results showed that JTKY treatment decreased the wet-to-dry ratio in the lung tissue, total protein concentration, and total cell count of the bronchoalveolar lavage fluid (BALF). Hematoxylin and Eosin (HE) and Masson staining indicated that the JTKY treatment alleviated the pathological changes and decreased the fibrotic contents in the lungs. JTKY treatment also decreased the expression of pro-inflammatory cytokines [interleukin (IL)-1β, IL-6, and tumor necrosis factor-alpha (TNF-α)] and increased the levels of immunomodulatory cytokines (IL-4 and IL-10) in the BALF and serum. Flow cytometry analysis showed that the JTKY treatment lowered the ratio of CD86+/CD206+ macrophages in the BALF, decreased inducible nitric oxide synthase (iNOS) level, and increased arginase 1 (Arg-1) level in lung. JTKY also lowered CD11b+Ly6G+ neutrophils in BALF and decreased myeloperoxidase (MPO) activity in lung. Moreover, it also elevated superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities and decreased methane dicarboxylic aldehyde (MDA) level in lung. Untargeted metabolomic analysis showed that the JTKY treatment could affect 19 metabolites in lung, such as L-adrenaline, L-asparagine, ornithine, and alpha-ketoglutaric acid. These metabolites are associated with the synthesis and degradation of ketone bodies, butanoate, alanine, aspartate, and glutamate metabolism, and tricarboxylic acid (TCA) cycle processes. In conclusion, our study demonstrated that treatment with JTKY ameliorated poly(I:C)-induced pneumonia. The mechanism of action of JTKY may be associated with the inhibition of the inflammatory response, the reduction of oxidative stress, and the regulation of the synthesis and degradation of ketone bodies, TCA cycle, and metabolism of alanine, aspartate, glutamate, and butanoate processes in lung.

Dissipative biocatalytic cascades and gated transient biocatalytic cascades driven by nucleic acid networks.

Living systems consist of complex transient cellular networks guiding structural, catalytic, and switchable functions driven by auxiliary triggers, such as chemical or light energy inputs. We introduce two different transient, dissipative, biocatalytic cascades, the coupled glucose oxidase (GOx)/horseradish peroxidase (HRP) glucose–driven oxidation of 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS2−) to the radical anion (ABTS•−) and the lactate dehydrogenase (LDH)/nicotinamide adenine dinucleotide (NAD+) lactate–driven reduction of NAD+ to NADH. The transient biocatalytic systems are driven by nucleic acid reaction modules using a nucleic acid fuel strand L1′ and a nicking enzyme, Nt.BbvCI, as fuel-degrading catalyst, leading to the dynamic spatiotemporal transient formation of structurally proximate biocatalysts activating the biocatalytic cascades and transient coupled processes, including the generation of chemiluminescence and the synthesis of alanine. Subjecting the mixture of biocatalysts to selective inhibitors allows the gated transient operation of the biocatalysts. The kinetics of transient biocatalytic cascades are accompanied by kinetic models and computational simulations.

Effect of Exogenous Melatonin Application on the Grain Yield and Antioxidant Capacity in Aromatic Rice under Combined Lead-Cadmium Stress.

This study aimed to determine the mechanism of exogenous melatonin application in alleviating the combined Pb and Cd (Pb-Cd) toxicity on aromatic rice (Oryza sativa L.). In this study, a pot experiment was conducted; two aromatic rice varieties, Yuxiangyouzhan and Xiangyaxiangzhan, were selected, and sprays using 50, 100, 200, and 400 μmol L−1 melatonin (denoted as S50, S100, S200, and S400) and irrigation using 100, 300, and 500 μmol L−1 melatonin (denoted as R100, R300, and R500) were also selected. The results showed that, under the S50, S100, and S200 treatments, the Pb content of aromatic rice grain decreased, and the grain yield increased significantly. Moreover, the application of exogenous melatonin significantly reduced the accumulation of H2O2 in rice leaves at maturity under Cd–Pb stress and reduced the MDA content in Xiangyaxiangzhan leaves. In addition, the microbial community structure changed significantly under S50 and R300 treatments. Some pathways, such as the synthesis of various amino acids and alanine, aspartate, and glutamate metabolism, were regulated by S50 treatment. Overall, melatonin application improved aromatic rice grain yield while reducing heavy metal accumulation by regulating the antioxidant capacity and metabolites in aromatic rice plants and altering the physicochemical properties and microbial community structures of the soil.

Transcriptome Analysis of Halotolerant Staphylococcus saprophyticus Isolated from Korean Fermented Shrimp.

Saeu-jeotgal, a Korean fermented shrimp food, is commonly used as an ingredient for making kimchi and other side dishes. The high salinity of the jeotgal contributes to its flavor and inhibits the growth of food spoilage microorganisms. Interestingly, Staphylococcus saprophyticus was discovered to be capable of growth even after treatment with 20% NaCl. To elucidate the tolerance mechanism, a genome-wide gene expression of S. saprophyticus against 0%, 10%, and 20% NaCl was investigated by RNA sequencing. A total of 831, 1314, and 1028 differentially expressed genes (DEGs) were identified in the 0% vs. 10%, 0% vs. 20%, and 10% vs. 20% NaCl comparisons, respectively. The Clusters of Orthologous Groups analysis revealed that the DEGs were involved in amino acid transport and metabolism, transcription, and inorganic ion transport and metabolism. The functional enrichment analysis showed that the expression of the genes encoding mechanosensitive ion channels, sodium/proton antiporters, and betaine/carnitine/choline transporter family proteins was downregulated, whereas the expression of the genes encoding universal stress proteins and enzymes for glutamate, glycine, and alanine synthesis was upregulated. Therefore, these findings suggest that the S. saprophyticus isolated from the saeu-jeotgal utilizes different molecular strategies for halotolerance, with glutamate as the key molecule.

Isotope effects at the origin of life: Fingerprints of the Strecker synthesis

Strecker synthesis creates α-amino acids from prebiotically plausible substrates (cyanide, ammonia, and aldehydes) and is widely hypothesized to be a key mechanism in the chemistry that led to life on Earth and on other planets. To better constrain the synthetic environments and precursors of abiotic α-amino acids, and to determine unique signatures of abiogenic amino acids, we measured the molecular-averaged and site-specific carbon and nitrogen isotope effects for the Strecker synthesis of alanine. The reaction steps of the Strecker synthesis can be divided into two groups: an initial series of reversible amination and nitrile-addition reactions (‘equilibration’) and a second series of irreversible hydrolysis reactions (‘hydrolysis’). The equilibration of cyanide, acetaldehyde, and ammonia with the intermediate, α-aminopropionitrile (α-APN), has a measured 55.1‰ equilibrium nitrogen isotope effect between the 15N-rich amine nitrogen in α-aminopropionitrile and the 15N-poor ammonia and a 20.0‰ equilibrium carbon isotope effect between the 13C-poor C-2 site in α-aminopropionitrile and the 13C-rich carbonyl carbon in acetaldehyde. The first irreversible hydrolysis step is inferred to have an up to 10‰ normal carbon fractionation (i.e., faster for 12C, slower for 13C) for the whole molecule, but it also has one or more side reactions that deplete the reactive α-APN reservoir by up to 15‰. The second hydrolysis step has a 15.4‰ normal kinetic isotope effect on the amide (C-1) site of alaninamide, which becomes the carboxyl site of alanine. Other α-amino acids will likely experience similar nitrogen isotope fractionations between ammonia and their amine sites, and similar carbon isotope fractionations between the carbonyl carbon in reactant aldehydes or ketones and the intermediate α-aminonitrile, and between cyanide and the carboxyl site. Therefore, these isotope effects allow us to predict the carbon and nitrogen isotopic contents and intramolecular structures of α-amino acids formed by Strecker synthesis based on their substrates’ isotopic compositions, or to infer the isotopic compositions of substrates from which amino acids formed, for example in the case of the amino-acid-rich carbonaceous chondrites. The site-specific C and N isotopic compositions of amino acids formed by Strecker chemistry contrast with those typical of terrestrial biosynthetic amino acids, so these data also provide a means of discriminating between biogenic and abiogenic α-amino acids.

Proteomic Shifts Reflecting Oxidative Stress and Reduced Capacity for Protein Synthesis, and Alterations to Mitochondrial Membranes in Neurospora crassa Lacking VDAC.

Voltage-dependent anion-selective channels (VDAC) maintain the bidirectional flow of small metabolites across the mitochondrial outer membrane and participate in the regulation of multiple cellular processes. To understand the roles of VDAC in cellular homeostasis, preliminary proteomic analyses of S100 cytosolic and mitochondria-enriched fractions from a VDAC-less Neurospora crassa strain (ΔPor-1) were performed. In the variant cells, less abundant proteins include subunits of translation initiation factor eIF-2, enzymes in the shikimate pathway leading to precursors of aromatic amino acids, and enzymes involved in sulfate assimilation and in the synthesis of methionine, cysteine, alanine, serine, and threonine. In contrast, some of the more abundant proteins are involved in electron flow, such as the α subunit of the electron transfer flavoprotein and lactate dehydrogenase, which is involved in one pathway leading to pyruvate synthesis. Increased levels of catalase and catalase activity support predicted increased levels of oxidative stress in ΔPor-1 cells, and higher levels of protein disulfide isomerase suggest activation of the unfolded protein response in the endoplasmic reticulum. ΔPor-1 cells are cold-sensitive, which led us to investigate the impact of the absence of VDAC on several mitochondrial membrane characteristics. Mitochondrial membranes in ΔPor-1 are more fluid than those of wild-type cells, the ratio of C18:1 to C18:3n3 acyl chains is reduced, and ergosterol levels are lower. In summary, these initial results indicate that VDAC-less N. crassa cells are characterized by a lower abundance of proteins involved in amino acid and protein synthesis and by increases in some associated with pyruvate metabolism and stress responses. Membrane lipids and hyphal morphology are also impacted by the absence of VDAC.