Amino Acids Isoleucine Research Articles

Nutritional Profile Comparison of Wild and Tagal Indigenous Mahseer (Tor duoronensis) from Borneo

Little is known about the nutritional status of mahseer (Tor duoronensis), a high-value indigenous freshwater species of Borneo. The present study evaluated the proximate, fatty acid, and amino acid compositions in carcasses of juvenile wild and conserved (tagal system) mahseer. The crude lipid content of the wild-caught fish was significantly higher (P < 0.05) than tagal fish. Mahseer from tagal system contained significantly higher (P < 0.05) fatty acid concentrations of C14:0, C15:0, C16:0, C16.1, C17:0, C18:0, C18:1 n9t, C20:0, C20:4n6, C20:5n3, and C22:6n3, and lower concentrations of C12:0, C18:1 n9c, C18:2 n6c, C18:3 n3, C20:1, and C21:0 fatty acids than wild mahseer. The percentages of total saturated fatty acids and n-3:n-6 ratio were higher in the tagal system than in wild mahseer. Tor duoronensisharvested from the tagal system showed higher total amino acid concentration (756.74 ± 225.21 ng g-1) compared to wild fish (652.10 ± 249.50 ng g-1). Among the amino acid recorded, cysteine content was high in both wild (147.12 ± 206.62 ng g-1) and tagal fish (164.37 ± 229.39 ng g-1). Nine essential amino acids (histidine, arginine, threonine, valine, methionine, lysine, isoleucine, and phenylalanine) that are important in fish’s metabolic and physiological responses were present in both fish groups. This study indicates that the biochemical composition of Tor duoronensis was significantly influenced by their habitat, and these differences may be explained by the components of the fish’s diet. The information generated from this study is beneficial for developing nutritionally balanced practical feed for this species.

Intracorneal iontophoretic delivery of triamcinolone acetonide prodrugs: Physicochemical parameters guiding electrotransport.

Intracorneal delivery of ten amino acid (alanine, arginine, asparagine, glutamine, glycine, histidine, isoleucine, lysine, methionine and valine) ester prodrugs of triamcinolone acetonide (TA-AA) was investigated in vitro, using a corneal iontophoresis device (IONTOFOR-CXL; SOOFT Italia S.p.A.) approved for clinical use in the treatment of keratoconus. Short duration iontophoresis (1mA for 5min) was performed and intracorneal deposition of TA was quantified by HPLC-UV and UHPLC-MS/MS. The data evidenced the clear advantage of TA-AA prodrug iontophoresis compared to passive delivery and revealed unexpected and prodrug dependent deposition profiles. Despite their superior electrical mobility, intracorneal delivery of dications, TA-Arg and TA-Lys, did not outperform that of TA-Ala and TA-Gly. In silico investigations to relate the TA-AA prodrugs' physicochemical properties to their electrotransport confirmed that increased lipophilicity potential did not favour iontophoretic transport. For TA-Ala and TA-Gly, it was hypothesized that the greater charge distribution and decreased tendency to interact with the corneal tissue via electrostatic and H-bonds contributed to their successful iontophoretic delivery. Intracorneal biodistribution of TA confirmed that TA-Gly iontophoresis resulted in supratherapeutic concentrations in deep corneal stroma, exceeding TA IC50 by∼104-fold. The results clearly demonstrated the successful combination of the clinically approved SOOFT iontophoretic device and the TA-AA prodrugs for targeted corneal iontophoretic delivery.

Valine Restriction Extends Survival in a Drosophila Model of Short-Chain Enoyl-CoA Hydratase 1 (ECHS1) Deficiency.

Short-chain enoyl-CoA hydratase 1 deficiency (ECHS1D) is a rare genetic disorder caused by biallelic pathogenic variants in the ECHS1 gene. ECHS1D is characterised by severe neurological and physical impairment that often leads to childhood mortality. Therapies such as protein and single nutrient-restricted diets show poor efficacy, whereas the development of new treatments is hindered by the low prevalence of the disorder and a lack of model systems for treatment testing. Here, we report on the establishment of a Drosophila model of ECHS1D. Flies carrying mutations in Echs1 (CG6543) were characterised for their physical and metabolic phenotypes, and dietary intervention to improve fly model health was explored. The Echs1 null larvae recapitulated human ECHS1D phenotypes including poor motor behaviour and early mortality and could be rescued by the expression of a human ECHS1 transgene. We observed that both restriction of valine in isolation, or all branched-chain amino acids (BCAAs-leucine, isoleucine and valine) together, extended larval survival, supporting the idea that reducing BCAA pathway catabolic flux is beneficial in this disorder. Further, metabolic profiling revealed substantial changes to carbohydrate metabolism, suggesting that Echs1 loss causes widespread metabolic dysregulation beyond valine metabolism. The similarities between Drosophila and human ECHS1D suggest that the fly model is a valuable animal system in which to explore mechanisms of pathogenesis and novel treatment options for this disorder.

Nutritional Potential of Maranthes polyandra (Benth.) Prance Fruit Kernels: A Comprehensive Analysis of Macronutrient, Mineral, and Phytochemical Profiles in Burkina Faso

Humans have been using plants in traditional medicine and for food security for millions of years. Today, several plants are being studied for their nutritional and medicinal value. The aim of this study was to determine the macronutrient composition, the characteristics of the oil and the mineral and phytochemical amino acid profiles of the fruit kernels of Maranthes polyandra (Benth.) Prance collected in the Cascades region of Burkina Faso. Macronutrient composition, oil characteristics and phytochemical profile were determined using standard methods. Amino acid and mineral profiles were determined by liquid chromatography coupled to a tandem mass spectrometer (LC-MS/MS) and inductively coupled plasma atomic emission spectrometry (ICP-AES) respectively. The results of the analyses revealed average levels of protein (14.5 ± 0.2 g/100g), lipids (55.5 ± 0.5 g/100g) and carbohydrates (25.2 g/100g). The average saponification, iodine, acid and peroxide values of the oils were 190.92 mg KOH/g oil, 148.11 g iodine/100 g oil, 0.24 mg KOH/g oil and 25.33 mEq of O2/kg oil. Essential amino acids (Threonine, Histidine, Isoleucine and Leucine) and non-essential amino acids (Alanine, Glycine, Arginine and Glutamic acid) were reported at varying levels. Almonds also contained ten types of minerals (B, Ca, Fe, K, Mg, Na, P, Si, Sr and Zn), total fiber (11.10%), phytate (12.46 g/kg) and polyphenols (0.13%). Almonds contained simple sugars (mg/kg) such as D-mannopyranose (41.80), Ribose (149.27), L-Rhamnose monohydrate (70.86), Glucuronic acid (161.41), Trigalacturonic acid (19.44), α-D-glucose (608.82), α-D-galactose (253.05), Aldehydo- D-xylose (22.23), Aldehydo-L-arabinose (49.72), Fucopyranose (510.58). Fructose was not detected. This study revealed that the fruit kernels of M. polyandra are highly rich in nutrients and can be used in food fortification to combat malnutrition in Burkina Faso.

Comprehensive analysis of lysine lactylation and its potential biological significance in clear cell renal cell carcinoma

BackgroundClear cell renal cell carcinoma (ccRCC) is a common histological subtype of malignant renal neoplasm. Protein lysine lactylation (Kla) plays a crucial role in tumor metabolic reprogramming. However, little is known regarding the distribution and potential biological functions of Kla in ccRCC. This study aimed to systematically investigate the role of Kla in ccRCC.MethodsA total of 12 ccRCC samples were collected from 6 patients. Western blotting was performed to determine the trend of Kla-modified proteins in ccRCC. Liquid chromatography–tandem mass spectrometry was used to quantitatively analyze Kla in ccRCC. Gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and protein–protein interaction (PPI) network analyses were conducted to clarify the biological functions and interactional relationships of differentially lactylated proteins (DLPs).ResultsIn total, 239 DLPs, including 441 lactylated sites, were identified by comparing ccRCC tissues with adjacent normal tissues. Kla-related enzymes have a higher affinity for alanine than for other amino acid residues in ccRCC. Subcellular localization analysis revealed that most DLPs were localized in the cytoplasm and mitochondria. GO enrichment analysis showed that most of the DLPs were enriched in metabolism-associated biological processes, including the purine ribonucleotide, monocarboxylic acid, ribonucleoside triphosphate, purine nucleoside triphosphate, and ATP metabolic processes. KEGG analysis indicated that most DLPs were also enriched in metabolism-related pathways, including glycolysis, amino acid (valine, leucine, and isoleucine) degradation, pyruvate metabolism, fatty acid degradation, and the citrate cycle. The top 20 hub proteins were screened from the PPI network based on their degree ranks.ConclusionsThis study revealed the role of Kla in ccRCC, which will extend our understanding of the potential molecular mechanisms underlying ccRCC formation and progression. These key Kla-modified proteins may be promising therapeutic targets for the treatment of ccRCC. However, further molecular experiments are required to validate these findings.

Integration of multi-omics layers empowers precision diagnosis through unveiling pathogenic mechanisms on maple syrup urine disease.

Maple syrup urine disease (MSUD) is a rare inherited metabolic disorder characterized by deficient activity of the branched-chain alpha-ketoacid dehydrogenase (BCKDH) complex, required to metabolize the amino acids leucine, isoleucine, and valine. Despite its profound metabolic implications, the molecular alterations underlying this metabolic impairment had not yet been completely elucidated. We performed a comprehensive multi-omics integration analysis, including genomic, epigenomic, and transcriptomic data from fibroblasts derived from a cohort of MSUD patients and unaffected controls to genetically characterize an MSUD case and to unravel the MSUD pathophysiology. MSUD patients exhibit a defined episignature that reshapes the global DNA methylation landscape, resulting in the stimulation of HOX cluster genes and the restriction of cell cycle gene-related signatures. Subsequent data integration revealed the impact of AP1-related and CEBPB transcription factors on the observed molecular reorganization, with MEIS1 emerging as a potential downstream candidate affected by robust epigenetic repression in MSUD patients. Furthermore, the integration of multi-omics layers facilitated the identification of a strong epigenetic repression in the DBT promoter in a patient wherein no BCKDH pathogenic variants had been detected. A Circular Chromatin Conformation Capture assay indicated a disturbance of the interactions of DBT promoter, thereby unveiling alternative modes of disease inheritance. Integration of multi-omics data unveiled underlying molecular networks rewired in MSUD patients and represents a powerful approach with diagnostic potential for rare genetic disorders with unknown genetic bases.

Impact of Amino Acid Supplementation on Aroma Development in Vidal Icewine: A Comparative Study of Saccharomyces and Non-Saccharomyces Fermentations

This study explored the effects of branched-chain amino acid (leucine, isoleucine, valine, and threonine) supplementation on the aroma development of Vidal icewine, fermented separately using commercial Saccharomyces cerevisiae (F33) and non-Saccharomyces (Hanseniaspora uvarum, Hu) yeast. The results indicated that Hu played a pivotal role in enhancing aromatic complexity by producing higher concentrations of esters, medium-chain fatty acids, and terpenes. Particularly, samples with L-isoleucine (1.0 g/L) and L-threonine (1.0 g/L) significantly increased key compounds such as beta-damascenone, isoamyl acetate, and phenethyl acetate, resulting in pronounced floral, fruity, and honey-like aromas. Conversely, during F33 fermentation, samples with added valine (1.0 g/L), threonine (1.4 g/L), and L-leucine (0.6 g/L) exhibited higher concentrations of higher alcohols and ethyl esters, crucial for delivering floral-fruity aromas. These findings suggest that precisely controlling the supply of amino acids during fermentation, through the selection of appropriate yeast strains and amino acid types, can effectively control the aromatic properties of icewine. This study offers new insights for improving icewine quality.

Maple syrup urine disease diagnosed in a resource-limited setting in an infant in Nepal: a case report

BackgroundMaple Syrup Urine Disease (MSUD) is a rare inherited disorder of metabolism, which manifests early in life in classical forms. Recurrent illness and exertion aggravate neurotoxicity. This case highlights MSUD diagnosed in association with COVID-19 complications from Nepal.Case presentationWe present a case of a 4-month-old child with a biochemical diagnosis of flared-up MSUD. Initially presenting with chief complaints of fever, noisy breathing, chest retraction, cough along with lethargy and poor feeding since the first week of life, the child also had developmental delay with feeble neck holding and absent social smile. The child was diagnosed with COVID-19 pneumonia and admitted in the Intensive Care Unit, requiring mechanical ventilation for 12 days. Despite the clinical resolution of pneumonia, the child had multiple episodes of generalized seizures and was sickly and frail. An incessant peculiar odor emanating from the child led to strong suspicion of metabolic disorder. Qualitative screening for amino acids (FeCl3 and 2,4-dinitrophenylhdrazine/DNPH) in urine and further gas chromatography-mass spectrometry revealed increased branched-chain amino acids(valine, leucine, and isoleucine). With dietary restrictions, the child was doing well. However, unfortunately, after 10 days of discharge, the child succumbed to death.ConclusionsThis case highlights the outpouring of hidden metabolic disorders with the onset of new diseases. It could have been detected and managed earlier with expedited neonatal screening and proper intervention.

Association of maternal blood metabolomics and gestational diabetes mellitus risk: a systematic review and meta-analysis.

Gestational diabetes mellitus (GDM) is a common complication of pregnancy that has short- and long-term adverse effects. Therefore, further exploration of the pathophysiology of GDM and related biomarkers is important. In this study, we performed a systematic review and meta-analysis to investigate the associations between metabolites in blood detected via metabolomics techniques and the risk of GDM and to identify possible biomarkers for predicting the occurrence of GDM. We retrieved case‒control and cohort studies of metabolomics and GDM published in PubMed, Embase, and Web of Science through March 29, 2024; extracted metabolite concentrations, odds ratios (ORs), or relative risks (RRs); and evaluated the integrated results with metabolites per-SD risk estimates and 95% CIs for GDM. We estimated the results via the random effects model and the inverse variance method. Our study is registered in PROSPERO (CRD42024539435). We included a total of 28 case‒control and cohort studies, including 17,370 subjects (4,372 GDM patients and 12,998 non-GDM subjects), and meta-analyzed 67 metabolites. Twenty-five of these metabolites were associated with GDM risk. Some amino acids (isoleucine, leucine, valine, alanine, aspartate, etc.), lipids (C16:0, C18:1n-9, C18:1n-7, lysophosphatidylcholine (LPC) (16:0), LPC (18:0), and palmitoylcarnitine), and carbohydrates and energy metabolites (glucose, pyruvate, lactate, 2-hydroxybutyrate, 3-hydroxybutyrate) were discovered to be associated with increased GDM risk (hazard ratio 1.06-2.77). Glutamine, histidine, C14:0, and sphingomyelin (SM) (34:1) were associated with lower GDM risk (hazard ratio 0.75-0.84). These findings suggest that these metabolites may play essential roles in GDM progression, and serve as biomarkers, contributing to the early diagnosis and prediction of GDM.

Late-life protein or isoleucine restriction impacts physiological and molecular signatures of aging.

Restricting the intake of protein or the branched-chain amino acid isoleucine promotes healthspan and extends lifespan in young or adult mice. However, their effects when initiated in aged animals are unknown. Here we investigate the consequences of consuming a diet with 67% reduction of all amino acids (low AA) or of isoleucine alone (low Ile), in male and female C57BL/6J.Nia mice starting at 20 months of age. Both dietary regimens effectively promote overall metabolic health without reducing calorie intake. Both low AA and low Ile diets improve aspects of frailty and slow multiple molecular indicators of aging rate; however, the low Ile diet reduces grip strength in both sexes and has mixed, sexually dimorphic effects on the heart. These results demonstrate that low AA and low Ile diets can promote aspects of healthy aging in aged mice and suggest that similar interventions might promote healthy aging in older adults.

Improvement of the Meat Quality of Holstein Bulls Fed a Diet Enriched with Oregano Oil.

Beef is a dominant meat commodity produced mainly from beef cattle breeds. Dairy cattle breeds are reared for milk production, but their calves could be raised to produce high quality meat subject to designated feeding. The objective here was to investigate the role of oregano oil-enriched diets on the quality characteristics of meat produced by Holstein bulls. Fifty Holstein bulls, aged 12 ± 2 months and with an average weight of 365 ± 60 kg, were used. They were equally allocated in two experimental groups (n = 25). One group (group CON) was offered a basal diet and the other (group OREG) the same diet enriched with 50 mg/kg DM of oregano oil, for a period of 90 days. Afterwards, meat samples were subjected to microbiological (1 h after slaughter), physicochemical (1st and 15th day after slaughter), and sensory analyses (15th day after slaughter). The beef of group OREG is characterized with improved color parameters and a lower malondialdehyde (MDA) production rate and content (68.52 ng/g versus 105.91 ng/g, p = 0.01). The sensory evaluation of beef revealed higher scores for group OREG in flavor (3.88 versus 3.17, p = 0.0007) and color (3.87 versus 3.21, p = 0.001), as well as higher nutritional value, which was associated with significantly higher levels of alanine, arginine, aspartic acid, histidine, isoleucine, leucine, lysine, valine, and total amino acids (p < 0.05). Thus, the results suggest that the quality of the meat of Holstein bulls can be improved when they are fed a diet enriched with oregano oil.

Acute metabolic decompensation after liver transplant in a patient with maple syrup urine disease.

Maple syrup urine disease (MSUD) is an inborn error of metabolism characterized by the accumulation of branched-chain amino acids (leucine, isoleucine, and valine) caused by a defect in the branched-chain alpha-keto acid dehydrogenase complex. Liver transplant is an effective therapy for MSUD, and patients can usually tolerate a regular diet after transplant without symptomatic metabolic decompensation. Most post-transplant patients do not follow a sick-day diet. We report a case of a 7-year-old male with MSUD Type IA, status post-liver transplant at 2 years of age, who presented with profound encephalopathy following poor oral intake and vomiting for 3 days. Broad laboratory workup was significant for hyperleucinosis and an unrevealing infectious workup. We conducted a review of eight post-liver transplant MSUD patients followed at Washington University in St. Louis. The review revealed that plasma amino acids were generally not checked during intercurrent illnesses in this patient cohort. While most of our patients have not had documented encephalopathy, one of the patients with epilepsy had a seizure during a gastrointestinal illness. Based on the review of the literature and from our center's experience, acute metabolic decompensation with intercurrent illnesses in MSUD patients after liver transplant appears to be rare. This case report raises awareness that patients with MSUD are still at risk of developing metabolic crisis post-liver transplant and provides additional insight into the risk factors associated with metabolic decompensation in this patient cohort.

Classification of colorectal cancer patients based on serum micronutrients: An exploratory investigation

BackgroundColorectal cancer (CRC) is a growing global health challenge with a multifactorial etiology encompassing genetic susceptibility, nutrition, and inflammation in the bowel. ObjectiveTo examine micronutrient status in CRC patients undergoing CRC resection. DesignWe performed a case-control study including 13 consecutive CRC patients and 10 healthy controls (CTRL) comparing the serum levels of 29 micronutrients, namely Copper, Zinc, Selenium, Chromium, Manganese, Carnitine, Choline, Inositol, Methylmalonic acid (MMA), Vitamin (Vit) B1, Vit B2, Vit B3, Vit B5, Vit B6, Vit C, Vit A, Vit D3, Vit E, Vit K1, Vit K2 and the amino acids Serine, Valine, Leucine, Isoleucine, Asparagine, Glutamine, Arginine, Citrulline and Cysteine. ResultsAfter considering the effect of age and sex, copper, arginine, and cysteine were increased, while zinc, selenium, chromium, Vit B1, Vit K1, and Vit A were decreased in CRC patients in comparison with CTRL. Zinc levels perfectly predicted the diagnosis of CRC, and were associated with lymph nodes (pN), of the pTNM staging. Copper levels in serum were strongly associated with the pathological pTNM staging of CRC. ConclusionThough this is a preliminary study that needs confirmation with a larger longitudinal cohort, our results show that serum micronutrients are linked to tumor growth, likely caused by increased demand from tumor tissue associated with an aberrant cell proliferation and changes in the antioxidant function.

Biochar enhances soil interactions and the initial development of sugarcane

The present study evaluated the effects of biochar derived from sugarcane straw on the physicochemical and biological properties of soil and the initial development of sugarcane. Microcosm and pot experiments were conducted over 60 days to monitor variables such as pH, water retention capacity, microbial activity, initial growth, and the metabolomic profile of the plant leaves. The results indicated that biochar increased the soil water retention capacity without significantly affecting the pH. The biochar significantly promoted root length and mass and favored the growth of the soil microbiota, as reflected by an increase in the amount of microbial biomass carbon. Metabolomic analysis of sugarcane leaves revealed that soil conditioning with biochar at application rates of 3 and 5% w/w led to increased concentrations of the amino acids isoleucine and valine, accompanied by a reduction in galactose, maltose, and glucose levels. Furthermore, biochar treatment resulted in a decrease in aconitic acid and an increase in acetic and succinic acid concentrations. These findings suggest that biochar may be a promising strategy for enhancing the productivity and sustainability of sugarcane cultivation.

Integrated Metabolome and Transcriptome Analysis Reveals the Key Bitter Substances of Pleioblastus amarus Shoots at Different Developmental Stages

Pleioblastus amarus (bitter bamboo) shoots are a traditional forest vegetable in China, renowned for its nutritional composition and associated health benefits. This study aimed to investigate the dynamic changes in nutritional quality and bitterness-related metabolites, as well as corresponding gene expression in P. amarus shoots at three distinct growth stages, specifically at heights of 10 cm (BT1), 20 cm (BT2), and 40 cm (BT3). The results showed that the content of soluble sugars decreased while the content of total phenols and flavonoids, which contribute to bitterness, increased during the growth of P. amarus shoots. In addition, comparative analysis of BT2 vs. BT1, BT3 vs. BT1, and BT3 vs. BT2 revealed 14, 43, and 36, respectively, differentially accumulated metabolites (DAMs). Notably, BT2 exhibited upregulation of flavonoids, such as apigenin, while bitter amino acids, including phenylalanine and arginine, showed a marked increase in BT3. Moreover, differential gene expression analysis revealed 3161, 13268, and 3742 differentially expressed genes (DEGs) in the BT2 vs. BT1, BT3 vs. BT1, and BT3 vs. BT2 comparisons, respectively. An integrated analysis of metabolomic and transcriptomic data indicated that the metabolites and genes associated with the biosynthesis pathways of phenylalanine, tyrosine, tryptophan, arginine, and the branched-chain amino acids valine, leucine, and isoleucine were significantly enriched during the growth of P. amarus shoots. The study indicates that early-stage (≤20 cm) P. amarus shoots are mainly bitter due to flavonoids like apigenin, while the bitterness in later-stage (around 40 cm) shoots is largely due to bitter amino acids such as phenylalanine, arginine, valine, and isoleucine. This research highlights key compounds and genes related to bitterness, providing a basis for future research on enhancing the nutritional value and flavor of P. amarus shoots.

Unveil the molecular recognition mechanisms of triazine haptens and monoclonal antibodies: Establishing ic‐ELISA methods for triazine herbicide analysis in tea

AbstractThis study introduced two triazine haptens (with 0 or 1 carbon connecting arms at nonisopropyl positions) and developed two heterologous indirect competitive enzyme‐linked immunosorbent assay (ic‐ELISA) methods. The IC50 values for the 17 triazine pesticides ranged from 0.84 ng/mL to 225.63 ng/mL, and the established method was applied to determine their concentrations in tea samples. This study aimed to address the current debate on the recognition mechanism of triazine haptens with shorter connecting arms. This study produced two new findings by investigating the recognition mechanism of triazine haptens and antibodies. On the one hand, the specific amino acid isoleucine (ILE‐219) is crucial for isopropyl recognition. On the other hand, the selection of the connecting site of the triazine hapten has a significant effect on the specificity of the antibody, with the influence of the length of the carbon connecting arm being secondary. Our newly designed haptens show promise for diverse triazine pesticide immunoassay applications. This discovery established a foundational framework for constructing a tailored antibody library for pollutants.

Comparative Metabolome and Transcriptome Analyses Reveal Molecular Mechanisms Involved in the Responses of Two Carex rigescens Varieties to Salt Stress.

Salt stress severely inhibits crop growth and production. The native turfgrass species Carex rigescens in northern China, exhibits extraordinary tolerance to multiple abiotic stresses. However, little is known about its specific metabolites and pathways under salt stress. To explore the molecular metabolic mechanisms under salt stress, we conducted metabolome analysis combined with transcriptome analysis of two varieties of Carex rigescens with differing salt tolerances: salt-sensitive Lvping NO.1 and salt-tolerant Lvping NO.2. After 5 days of salt treatment, 114 and 131 differentially abundant metabolites (DAMs) were found in Lvping NO.1 and Lvping NO.2, respectively. Among them, six amino acids involved in the amino acid biosynthesis pathway, namely, valine, phenylalanine, isoleucine, tryptophan, threonine, and serine, were accumulated after treatment. Furthermore, most DAMs related to phenylalanine biosynthesis, metabolism, and phenylpropanoid biosynthesis increased under salt stress in both varieties. The expression profiles of metabolism-associated genes were consistent with the metabolic profiles. However, genes including HCT, β-glucosidases, and F5H, and metabolite 4-hydroxycinnamic acid, of the two varieties may account for the differences in salt tolerance. Our study provides new insights into the mechanisms underlying salt tolerance in Carex rigescens and reveals potential metabolites and genes to improve crop resilience to saline environments.

Metabolic engineering of the serine/glycine network as a means to improve the nitrogen content of crops.

In plants, L-serine (Ser) biosynthesis occurs through various pathways and is highly dependent on the atmospheric CO2 concentration, especially in C3 species, due to the association of the Glycolate Pathway of Ser Biosynthesis (GPSB) with photorespiration. Characterization of a second plant Ser pathway, the Phosphorylated Pathway of Ser Biosynthesis (PPSB), revealed that it is at the crossroads of carbon, nitrogen, and sulphur metabolism. The PPSB comprises three sequential reactions catalysed by 3-phosphoglycerate dehydrogenase (PGDH), 3-phosphoSer aminotransferase (PSAT) and 3-phosphoSer phosphatase (PSP). PPSB was overexpressed in plants exhibiting two different modes of photosynthesis: Arabidopsis (C3 metabolism), and maize (C4 metabolism), under ambient (aCO2) and elevated (eCO2) CO2 growth conditions. Overexpression in Arabidopsis of the PGDH1 gene alone or PGDH1, PSAT1 and PSP1 in combination increased the Ser levels but also the essential amino acids threonine (aCO2), isoleucine, leucine, lysine, phenylalanine, threonine and methionine (eCO2) compared to the wild-type. These increases translated into higher protein levels. Likewise, starch levels were also increased in the PPSB-overexpressing lines. In maize, PPSB-deficient lines were obtained by targeting PSP1 using Cas9 endonuclease. We concluded that the expression of PPSB in maize male gametophyte is required for viable pollen development. Maize lines overexpressing the AtPGDH1 gene only displayed higher protein levels but not starch at both aCO2 and eCO2 conditions, this translated into a significant rise in the nitrogen/carbon ratio. These results suggest that metabolic engineering of PPSB in crops could enhance nitrogen content, particularly under upcoming eCO2 conditions where the activity of GPSB is limited.

An easy and sensitive assay for acetohydroxyacid synthases based on the simultaneous detection of substrates and products in a single step

Acetohydroxyacid synthase (AHAS, EC 2.2.1.6) catalyzes the first step in the synthesis of the branched-chain amino acids valine, leucine, and isoleucine, pathways being present in plants and microorganisms, but not in animals. Thus, AHAS is an important target for numerous herbicides and, more recently, for the development of antimicrobial agents. The need to develop new and optimized herbicides and pharmaceuticals requires a detailed understanding of the biochemistry of AHAS. AHAS transfers an activated two-carbon moiety derived from pyruvate to either pyruvate or 2-oxobutyrate as acceptor substrates, forming 2-acetolactate or 2-acetohydroxy-2-butyrate, respectively. Various methods have been described in the literature to biochemically characterize AHAS with respect to substrate preferences, substrate specificity, or kinetic parameters. However, the simultaneous detection and quantification of substrates and unstable products of the AHAS-catalyzed reaction have always been a challenge. Using AHAS isoform II from Escherichia coli, we have developed a sensitive assay for AHAS-catalyzed reactions that uses derivatization with ethyl chloroformate to stabilize and volatilize all reactants in the aqueous solution and detect them by gas chromatography coupled to flame ionization detection or mass spectrometry. This assay allows us to characterize the product formation in reactions in single and dual substrate reactions and the substrate specificity of AHAS, and to reinterpret previous biochemical observations. This assay is not limited to the AHAS-catalyzed reactions, but should be applicable to studies of many metabolic pathways.Graphical

Amino Acid Residue-Specific Ramachandran Distributions Derived from a Simple Mean Field Potential.

Protein dynamics in the unfolded state, in the context of early stage protein folding or intrinsically disordered proteins (IDPs), is not well understood. The discovery of IDPs, and their sequence-dependent dynamics, has led to many computational and experimental investigations regarding the conformational preferences of short oligopeptides and individual amino acid residues in the unfolded state. As proteins consist of sequences of amino acid residues, characterizing the intrinsic conformational preferences of the individual residues in the unfolded state is crucial for understanding the emergent conformations of peptides and proteins. While advances have been made in understanding conformational preferences, the atomistic mechanisms driving these preferences remain unresolved. In this work, we show that the distributions of atomic overlaps between backbone and side chain atoms in Ramachandran space are unique for amino acid residue mimetic structures alanine, valine, leucine, and isoleucine in Ramachandran space indicating unique intrapeptide energy landscapes for each residue. We then construct a mean field potential consisting of only an empirical peptide backbone-water and average intrapeptide Lennard-Jones contributions to explore their influence on the conformational preferences. With this fairly simple model, we were able to produce Ramachandran distributions that qualitatively agree with previously reported experimental and computational predictions about the conformational preferences of these amino acid residues in the unfolded state in water. Our results indicate these conformational preferences are the result of the balance between pPII-stabilizing backbone-water interactions and repulsive side chain-backbone interactions where the latter will depend uniquely on the atomic makeup and geometry of the side chain.