Aminoadipic Acid Research Articles

A metabolomic analysis of the sex‐dependent Hispanic Paradox

In Mexican Americans, metabolic conditions, such as obesity and type 2 diabetes (T2DM), are not necessarily associated with an increase in mortality, which is the so-called Hispanic Paradox. In this cross-sectional analysis, we used a metabolomic analysis to look at the mechanisms behind the Hispanic Paradox. To do this, we examined dietary intake and body mass index (BMI; kg/m2) in men and women and their effects on serum metabolomic fingerprints in 70 Mexican Americans (26 men, 44 women). Although having different BMI values, the participants had many similar anthropometric and biochemical parameters such as systolic and diastolic blood pressure, total cholesterol, and LDL cholesterol, which supports the Paradox in these subjects. Plasma metabolomic phenotypes were measured using liquid chromatography tandem mass spectrometry (LC-MS/MS). A two-way ANOVA assessing sex, BMI, and the metabolome revealed 23 significant metabolites, such as 2-pyrrolidinone (P = 0.007), TMAO (P = 0.014), 2-aminoadipic acid (P = 0.019), and kynurenine (P = 0.032). Pathway and enrichment analyses discovered several significant metabolic pathways between men and women, including lysine degradation, tyrosine metabolism, and branch-chained amino acid (BCAA) degradation and biosynthesis. A log-transformed OPLS-DA model was employed and demonstrated a difference due to BMI in the metabolomes of both sexes. When stratified for caloric intake (< 2200 kcal/d vs > 2200 kcal/d), a separate OPLS-DA model showed clear separation in men, while females remained relatively unchanged. After accounting for caloric intake and BMI status, the female metabolome showed substantial resistance to alteration. Therefore, we provide a better understanding of the Mexican American metabolome that may help demonstrate how this population, particularly women, possesses a longer life expectancy despite several comorbidities and reveal the underlying mechanisms of the Hispanic Paradox.

The effect of combaine administration of L-arginine and omega-3 polyunsaturated fatty acids on the spectrum of free amino acids and biogen amines in hyppocampus of rats undergoing subtotal cerebral ischemia

The aim of this study was to estimate the changes in the pool of free amino acids and their derivatives in hippocampus of rats undergoing subtotal cerebral ischemia (SCI) and treated with L-arginine and omega-3 polyunsaturated fatty acids (PUFA). Experiment was held on 18 rats: 12 animals were undergoing bilateral filament occlusion of arteries carotid, 6 of them L-arginine and omega-3 PUFA was administrated. The drug omega-3 PUFA "Omegamed" (at a dose of 5 g/kg of body weight) was injected intragastrically during the week preceding the simulation of SIMG. L-arginine (at a dose of 100 mg/kg body weight) was injected intravenously just before ligation of the common carotid arteries. The analyses of free amino acids and their derivates levels in the blood plasma extracts were carried out by reversed phase HPLC. In the hippocampus of rats with SIGM, there was an increase in the levels of histidine, 3-methylhistidine, glutamine, α--aminobutyrate, isoleucine, leucine, valine, as well as a decrease in the levels of threonine, tyrosine, and α--aminoadipic acid. Administration of L-arginine and omega-3 PUFAs prevented ischemia-induced disruption of threonine, histidine, glutamine, α--aminobutyrate, α--aminoadipic acid levels, and also had a corrective effect on the serotonin system of the hippocampus.

Pharmacophore Modelling of Vanillin Derivatives, Favipiravir, Chloroquine, Hydroxychloroquine, Monolaurin and Tetrodotoxin as MPro inhibitors of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2)

Ligand-based pharmacophore modelling approach using four established antiviral drugs, namely remdesivir, lopinavir, ritonavir and hydroxychloroquine were analysed for COVID-19 inhibitors as training sets. Twenty vanillin derivatives together with monolaurin were used as test sets to evaluate potential as SARS-CoV-2 inhibitors. Structure-based pharmacophore modelling approach was also performed using Protein Data Bank information: PDB-5RE6, 5REX and 5RFZ in order to analyse the binding site and ligand-protein complex interactions. The pharmacophore modelling mode of 5RE6 displayed two Hydrogen Bond Acceptors (HBA) and one Hydrophobic (HY) interaction. Besides, the pharmacophore model of 5REX showed two HBA and two HY interactions. Finally, the pharmacophore model of 5RFZ showed three HBA and one HY interaction. Based on ligand-based approach, 20 Schiff-based vanillin derivatives, namely vanillin associated with methyl-6- aminopyridine-3-carboxylate (1), sepiapterin (2), 6-aminopyridine-3-carboxylic acid (3), 6-aminopyridine-2-carboxylic acid (4), pemoline (5), α-phenylglycine (6), 2-amino-4-hydroxy-3-methylpentanoic acid (7), 4-hydroxyphenylglycine (8), β-homoserine (9), allylglycine (10), oxamic acid (11) benzophenone hydrazine (12), 2-aminoadipic acid (13), D-alanyl-D-alanine (14), p-bromophenylalanine (15), nicotinic hydrazide (16), 4-hydroxybenzhydrazide (17), benzohydrazide (18), isonicotinic hydrazide (19), and phenylhydrazine (20) showed strong MPro inhibition activity. This was due to their good alignment and common features to PDB-5RE6. Similarly, monolaurin and tetrodotoxin displayed some significant activity against SARS-CoV-2. From structure-based approach, vanillin derivatives (1) to (12) displayed some potent MPro inhibition against SARS-CoV-2. Favipiravir, chloroquine and hydroxychloroquine also showed some significant MPro inhibition. Favipiravir showed good alignment and common pharmacophore features to PDB- 5RFZ, whereas chloroquine and hydroxychloroquine showed good alignment and common pharmacophore features to PDB-5REX.

Plasma levels of amino acids and derivatives in retinopathy of prematurity.

Background: Retinopathy of prematurity (ROP) is a retinal disease that causes blindness in premature infants. This study aimed to reveal the changes in amino acids and derivatives in the plasma of ROP patients compared with premature infants without ROP.Methods: Metabolomics targeting amino acids and their derivatives was conducted to assess their plasma levels in ROP patients (n=58) and premature infants without ROP (n=25), and KEGG pathway analysis was used to identify the involved pathways.Results: Among the 31 assessed metabolites, the levels of 4 amino acids were significantly altered in the ROP group. Creatinine was downregulated in the plasma of the ROP patients, while the levels of citrulline, arginine, and aminoadipic acid were upregulated in the ROP group. Significant correlations were identified between the ROP stage and plasma levels of citrulline, creatinine, and aminoadipic acid. The involved pathways included biosynthesis of amino acids, arginine and proline metabolism, and arginine biosynthesis.Conclusion: The plasma levels of citrulline, creatinine, arginine, and aminoadipic acid were significantly changed in ROP patients. These metabolites could be considered potential biomarkers of ROP, and their related metabolic pathways might be involved in ROP pathogenesis.

Preferential accumulation of the active S-(+) isomer in murine retina highlights novel mechanisms of vigabatrin-associated retinal toxicity

((S)-(+)/(R)-(-)) vigabatrin (SabrilR; γ-vinyl GABA), an antiepileptic irreversibly inactivating GABA-transaminase, was administered to male C57Bl6 J mice via continuous infusion (0, 40, 80 mg/kg/d) for 12 days. Our study design pooled retina, eye (minus retina), whole brain and plasma from n = 24 animals for each dose to provide n = 8 triplicates per treatment group. Hypothesizing that (S)-(+) VGB (active isomer) would preferentially accumulate in retina, we determined VGB isomers, comprehensive amino acids, and pharmacokinetic parameters. In brain, eye and plasma, the ((S)-(+)/(R)-(-)) ratio varied from 0.73 to 1.29 and 13.3 in retina, accompanied by a partition coefficient (tissue/plasma, ((S)-(+);(R)-(-))) of 5.8;0.34, 0.63;0.49, and 0.51;0.34 in retina, eye and brain, respectively. Racemic VGB (nmol/g; plasma, nmol/mL, range of means for dose) content was: retina, 25–36; eye (minus retina), 4.8–8.0; brain, 3.1–6.8 and plasma, 8.7−14.9. GABA tissue content (nmol/g) was 1246–3335, 18–64 and 2615–3200 as a function of VGB dose for retina, eye (minus retina) and brain, respectively. The retinal glial cell toxin 2-aminoadipic acid also increased with VGB dose (76−96 nmol/g). Partitioning of active (S)-(+) VGB to retina suggests the involvement of a stereospecific transporter, the identification of which could reveal new therapeutic paradigms that might mitigate VGB’s well-known retinal toxicity and expand its clinical utility.

Discovery and validation of an NMR-based metabolomic profile in urine as TB biomarker

Despite efforts to improve tuberculosis (TB) detection, limitations in access, quality and timeliness of diagnostic services in low- and middle-income countries are challenging for current TB diagnostics. This study aimed to identify and characterise a metabolic profile of TB in urine by high-field nuclear magnetic resonance (NMR) spectrometry and assess whether the TB metabolic profile is also detected by a low-field benchtop NMR spectrometer. We included 189 patients with tuberculosis, 42 patients with pneumococcal pneumonia, 61 individuals infected with latent tuberculosis and 40 uninfected individuals. We acquired the urine spectra from high and low-field NMR. We characterised a TB metabolic fingerprint from the Principal Component Analysis. We developed a classification model from the Partial Least Squares-Discriminant Analysis and evaluated its performance. We identified a metabolic fingerprint of 31 chemical shift regions assigned to eight metabolites (aminoadipic acid, citrate, creatine, creatinine, glucose, mannitol, phenylalanine, and hippurate). The model developed using low-field NMR urine spectra correctly classified 87.32%, 85.21% and 100% of the TB patients compared to pneumococcal pneumonia patients, LTBI and uninfected individuals, respectively. The model validation correctly classified 84.10% of the TB patients. We have identified and characterised a metabolic profile of TB in urine from a high-field NMR spectrometer and have also detected it using a low-field benchtop NMR spectrometer. The models developed from the metabolic profile of TB identified by both NMR technologies were able to discriminate TB patients from the rest of the study groups and the results were not influenced by anti-TB treatment or TB location. This provides a new approach in the search for possible biomarkers for the diagnosis of TB.

Metabolite biomarkers of type 2 diabetes mellitus and pre-diabetes: a systematic review and meta-analysis

BackgroundWe aimed to explore metabolite biomarkers that could be used to identify pre-diabetes and type 2 diabetes mellitus (T2DM) using systematic review and meta-analysis.MethodsFour databases, the Cochrane Library, EMBASE, PubMed and Scopus were selected. A random effect model and a fixed effect model were applied to the results of forest plot analyses to determine the standardized mean difference (SMD) and 95% confidence interval (95% CI) for each metabolite. The SMD for every metabolite was then converted into an odds ratio to create an metabolite biomarker profile.ResultsTwenty-four independent studies reported data from 14,131 healthy individuals and 3499 patients with T2DM, and 14 included studies reported 4844 healthy controls and a total of 2139 pre-diabetes patients. In the serum and plasma of patients with T2DM, compared with the healthy participants, the concentrations of valine, leucine, isoleucine, proline, tyrosine, lysine and glutamate were higher and that of glycine was lower. The concentrations of isoleucine, alanine, proline, glutamate, palmitic acid, 2-aminoadipic acid and lysine were higher and those of glycine, serine, and citrulline were lower in prediabetic patients. Metabolite biomarkers of T2DM and pre-diabetes revealed that the levels of alanine, glutamate and palmitic acid (C16:0) were significantly different in T2DM and pre-diabetes.ConclusionsQuantified multiple metabolite biomarkers may reflect the different status of pre-diabetes and T2DM, and could provide an important reference for clinical diagnosis and treatment of pre-diabetes and T2DM.

Health risks of chlorothalonil, carbendazim, prochloraz, their binary and ternary mixtures on embryonic and larval zebrafish based on metabolomics analysis

Chlorothalonil (CTL), carbendazim (CBZ), and prochloraz (PCZ) are fungicides widely used in many countries. The use of these fungicides raises concerns because they are often applied together or used in the same agricultural area. However, the toxicity of these fungicides or mixtures, especially to aquatic organisms, has received limited attention. Here, embryonic and larval zebrafish were exposed to indicated concentrations of CTL, CBZ, and PCZ and their binary (CTL+CBZ, CTL+PCZ and CBZ+PCZ) and ternary (CTL+CBZ+PCZ) mixtures for 24 h. Based on metabolomics analysis, we observed that hundreds of metabolites were altered, and glycolysis metabolism and amino acid metabolism were the two most affected pathways. Interestingly, a total of 9 and 26 metabolites changed significantly in embryos and larvae treated with all fungicides, respectively. Among these altered metabolites, 2-aminoadipic acid (2-AAA) levels increased significantly in all groups, indicating that 2-AAA potentially represents a useful biomarker for evaluating the toxicity of fungicides. Furthermore, the joint effects of CTL+PCZ on embryos and larvae, especially on amino acid metabolism, were weaker than those in other groups, but combined treatment did not influence individual fungicidal activity. Data acquired from metabolomics provided important insight for understanding the mechanism by which fungicides or their mixtures affect zebrafish.

Review of Lysine Metabolism with a Focus on Humans

Lysine cannot be synthesized by most higher organisms and, therefore, is an indispensable amino acid (IAA) that must be consumed in adequate amounts to maintain protein synthesis. Although lysine is an abundant amino acid in body proteins, lysine is limited in abundance in many important food sources (e.g. grains). Older observations assigned importance to lysine because animals fed a lysine-deficient diet did not lose weight as fast as animals placed upon other IAA-deficient diets, leading to the theory that there may be a special pool of lysine or metabolites that could be converted to lysine. The first step in the lysine catabolic pathway is the formation of saccharopine and then 2-aminoadipic acid, processes that are mitochondrial. The catabolism of 2-aminoadipic acid proceeds via decarboxylation to a series of CoA esters ending in acetyl-CoA. In mammals, the liver appears to be the primary site of lysine catabolism. In humans, the metabolic and oxidative response of lysine to diets either restricted in protein or in lysine is consistent with what has been measured for other IAAs with isotopically labeled tracers. Intestinal microflora are known to metabolize urea to ammonia and scavenge nitrogen (N) for the synthesis of amino acids. Studies feeding 15N-ammonium chloride or 15N-urea to animals and to humans, demonstrate the appearance of 15N-lysine in gut microbial lysine and in host lysine. However, the amount of 15N-lysine transferred to the host is difficult to assess directly using current methods. It is important to understand the role of the gut microflora in human lysine metabolism, especially in conditions where dietary lysine intake may be limited, but better methods need to be devised.

Prognostic Role of Serum Amino Acids in Head and Neck Cancer.

Introduction Serum amino acid (AA) profiles represent a valuable tool in the metabolic assessment of cancer patients; still, information on the AA pattern in head and neck cancer (HNC) patients is insufficient. The aim of the study was to assess whether serum AA levels were associated with the stage of neoplastic disease and prognosis in primary HNC patients. Methods Two hundred and two primary HNC patients were included in the study. Thirty-one AAs and derivatives were measured in serum through an ultraperformance liquid chromatography-mass spectrometry (UPLC-MS). The association between AA concentrations and the stage (advanced versus early) of HNC was estimated using a multivariable logistic regression model. A multivariable Cox regression model was used to evaluate the prognostic significance of each AA. Results At the multivariable logistic regression analysis, increased levels of alpha-aminobutyric acid, aminoadipic acid, histidine, proline, and tryptophan were associated with a reduced risk of advanced stage HNC, while high levels of beta-alanine, beta-aminobutyric acid, ethanolamine, glycine, isoleucine, 4-hydroxyproline, and phenylalanine were associated with an increased risk of advanced stage HNC. Furthermore, at multivariate analysis, increased levels of alpha-aminobutyric acid were associated with increased overall survival (OS), while high levels of arginine, ethanolamine, glycine, histidine, isoleucine, 4-hydroxyproline, leucine, lysine, 3-methylhistidine, phenylalanine, and serine were associated with decreased OS. Conclusions Our study suggests that AA levels are associated with the stage of disease and prognosis in patients with HNC. More study is necessary to evaluate if serum AA levels may be considered a hallmark of HNC and prove to be clinically useful markers of disease status and prognosis in HNC patients.

Gas explosion-induced acute blast lung injury assessment and biomarker identification by a LC-MS-based serum metabolomics analysis.

The objective of this study was to evaluate the histopathological effect of gas explosion on rats, and to explore the metabolic alterations associated with gas explosion-induced acute blast lung injury (ABLI) in real roadway environment using metabolomics analyses. All rats were exposed to the gas explosion source at different distance points (160 m and 240 m) except the control group. Respiratory function indexes were monitored and lung tissue analysis was performed to correlate histopathological effect to serum metabolomics. Their sera samples were collected to measure the metabolic alterations by ultra-performance liquid chromatography-mass spectrometry (UPLC-MS). HE staining in lung showed that the gas explosion caused obvious inflammatory pulmonary injury, which was consistent with respiratory function monitoring results and the serum metabolomics analysis results. The metabolomics identified 9 significantly metabolites different between the control- and ABLI rats. 2-aminoadipic acid, L-methionine, L-alanine, L-lysine, L-threonine, cholic acid and L-histidine were significantly increased in the exposed groups. Citric acid and aconitic acid were significantly decreased after exposure. Pathway analyses identified 8 perturbed metabolic pathways, which provided novel potential mechanisms for the gas explosion-induced ABLI. Therefore, metabolomics analysis identified both known and unknown alterations in circulating biomarkers, adding an integral mechanistic insight into the gas explosion-induced ABLI in real roadway environment.

An integrated omics approach to investigate summer mortality of New Zealand Greenshell™ mussels.

Green-lipped mussels, commercially known as Greenshell™ mussels (Perna canaliculus Gmelin 1791), contribute > $300 million to New Zealand's aquaculture exports. However, mortalities during summer months and potential pathogenic outbreaks threaten the industry. Thermal stress mechanisms and immunological responses to pathogen infections need to be understood to develop health assessment strategies and early warning systems. P. canaliculus were collected during a mortality event at a commercial aquaculture farm in Firth of Thames, New Zealand. Gill tissues from six healthy and six unhealthy mussels were excised and processed for metabolomic (GC-MS) and label-free proteomic (LC-MS) profiling. Univariate analyses were conducted separately on each data layer, with data being integrated via sparse multiple discriminative canonical correlation analysis. Pathway enrichment analysis was used to probe coordinated changes in functionally related metabolite sets. Findings revealed disruptions of the tricarboxylic acid (TCA) cycle and fatty acid metabolism in unhealthy mussels. Metabolomics analyses also indicated oxidative stress in unhealthy mussels. Proteomics analyses identified under-expression of proteins associated with cytoskeleton structure and regulation of cilia/flagellum in gill tissues of unhealthy mussels. Integrated omics revealed a positive correlation between Annexin A4 and CCDC 150 and saturated fatty acids, as well as a negative correlation between 2-aminoadipic acid and multiple cytoskeletal proteins. Our study demonstrates the ability of using integrative omics to reveal metabolic perturbations and protein structural changes in the gill tissues of stressed P. canaliculus and provides new insight into metabolite and protein interactions associated with incidences of summer mortality in this species.

Optimizing Charge Switching in Membrane Lytic Peptides for Endosomal Release of Biomacromolecules

Endocytic pathways are practical routes for the intracellular delivery of biomacromolecules. Along with this, effective strategies for endosomal cargo release into the cytosol are desired to achieve successful delivery. Focusing on compositional differences between the cell and endosomal membranes and the pH decrease within endosomes, we designed the lipid-sensitive and pH-responsive endosome-lytic peptide HAad. This peptide contains aminoadipic acid (Aad) residues, which serve as a safety catch for preferential permeabilization of endosomal membranes over cell membranes, and His-to-Ala substitutions enhance the endosomolytic activity. The ability of HAad to destabilize endosomal membranes was supported by model studies using large unilamellar vesicles (LUVs) and by increased intracellular delivery of biomacromolecules (including antibodies) into live cells. Cerebral ventricle injection of Cre recombinase with HAad led to Cre/loxP recombination in a mouse model, thus demonstrating potential applicability of HAad in vivo.

Optimizing Charge Switching in Membrane Lytic Peptides for Endosomal Release of Biomacromolecules

AbstractEndocytic pathways are practical routes for the intracellular delivery of biomacromolecules. Along with this, effective strategies for endosomal cargo release into the cytosol are desired to achieve successful delivery. Focusing on compositional differences between the cell and endosomal membranes and the pH decrease within endosomes, we designed the lipid‐sensitive and pH‐responsive endosome‐lytic peptide HAad. This peptide contains aminoadipic acid (Aad) residues, which serve as a safety catch for preferential permeabilization of endosomal membranes over cell membranes, and His‐to‐Ala substitutions enhance the endosomolytic activity. The ability of HAad to destabilize endosomal membranes was supported by model studies using large unilamellar vesicles (LUVs) and by increased intracellular delivery of biomacromolecules (including antibodies) into live cells. Cerebral ventricle injection of Cre recombinase with HAad led to Cre/loxP recombination in a mouse model, thus demonstrating potential applicability of HAad in vivo.

Changes in milk metabolome during the lactation of dairy cows based on 1H NMR and UHPLC–QToF/MS

The abundances of low-molecular-weight metabolites in milk have been known to change during the lactation stage, and this is of relevance to the milk nutritional or processing properties. Metabolomics was used to investigate changes in 46 milk samples from 19 Jersey cows. Metabolites were measured using both nuclear magnetic resonance and mass spectrometry based methods. Metabolites exhibiting change were identified with the aid of multivariate and univariate data analysis. The changing metabolites included amino acids and derivatives (taurine, purine, indoleacetic acid, aminoadipic acid), organic acids (uric acid, hippuric acid, citric acid), carbohydrates (N-acetyl-d-glucosamine, galactose-1-phosphate), vitamins (choline and pantothenic acid), nucleic acid-related compounds (cytosine, uracil and methylguanine) and fatty acids (lauric acid). This research not only provides a full understanding of the milk metabolomic changes due to the cow lactation stage but also can be used as the foundation for marker selection for further studies utilising milk metabolomics.

Noxious effects of selected food-occurring oxidized amino acids on differentiated CACO-2 intestinal human cells

The harmful effects of food-occurring oxidized amino acids, namely, aminoadipic acid (AAA), dityrosine (DTYR), L-kynurenine (KN), kynurenic acid (KA) and 3-nitrotyrosine (3NT), were studied on differentiated CACO-2 cells by flow cytometry and quantification of glutathione (GSH), and allysine. Cells were exposed to food-relevant doses (200 μM) of each compound for 4 or 72h and compared to a control (no stimulated cells). All oxidized amino acids induced apoptosis and results indicated that underlying mechanisms depended on the chemical nature of the species. AAA, KN and KA caused ROS generation and severe oxidative stress in 96%, 98% and 89% of exposed cells (77% in control cells), leading to significant GSH depletion and allysine accretion (1.5, 1.5 and 1.6 nmol allysine/mg protein, respectively at 4h; control: 0.22 nmol/mg protein; p < 0.05). DTYR and 3NT induced significant apoptosis to 29% and 25% of cells (control: 16%; p < 0.05) and necrosis to 28% and 26% of cells (control: 23%) at 72h by ROS-independent mechanisms. KN and KA were found to induce a cycle arrest effect on CACO-2 cells. These findings emphasize the potential harmful effects of the intake of oxidized proteins and amino acids and urge the necessity of carrying out further molecular studies.

The lysine derivative aminoadipic acid, a biomarker of protein oxidation and diabetes-risk, induces production of reactive oxygen species and impairs trypsin secretion in mouse pancreatic acinar cells

We have examined the effects of α-aminoadipic acid, an oxidized derivative from the amino acid lysine, on the physiology of mouse pancreatic acinar cells. Changes in intracellular free-Ca2+ concentration, the generation of reactive oxygen species, the levels of carbonyls and thiobarbituric-reactive substances, cellular metabolic activity and trypsin secretion were studied. Stimulation of mouse pancreatic cells with cholecystokinin (1 nM) evoked a transient increase in [Ca2+]i. In the presence of α-amoniadipic acid increases in [Ca2+]i were observed. In the presence of the compound, cholecystokinin induced a Ca2+ response that was smaller compared with that observed when cholecystokinin was applied alone. Stimulation of cells with cholecystokinin in the absence of Ca2+ in the extracellular medium abolished further mobilization of Ca2+ by α-aminoadipic acid. In addition, potential pro-oxidant conditions, reflected as increases in ROS generation, oxidation of proteins and lipids, were noted in the presence of α-aminoadipic acid. Finally, the compound impaired trypsin secretion induced by the secretagogue cholecystokinin. We conclude that the oxidized derivative from the amino acid lysine induces pro-oxidative conditions and the impairment of enzyme secretion in pancreatic acinar cells. α-aminoadipic acid thus creates a situation that could potentially lead to disorders in the physiology of the pancreas.

Structure-function investigation of 3-methylaspartate ammonia lyase reveals substrate molecular determinants for the deamination reaction.

The enzymatic reactions leading to the deamination of β-lysine, lysine, or 2-aminoadipic acid are of great interest for the metabolic conversion of lysine to adipic acid. Enzymes able to carry out these reactions are not known, however ammonia lyases (EC 4.3.1.-) perform deamination on a wide range of substrates. We have studied 3-methylaspartate ammonia lyase (MAL, EC 4.3.1.2) as a potential candidate for protein engineering to enable deamination towards β-lysine, that we have shown to be a competitive inhibitor of MAL. We have characterized MAL activity, binding and inhibition properties on six different compounds that would allow to define the molecular determinants necessary for MAL to deaminate our substrate of interest. Docking calculations showed that β-lysine as well as the other compounds investigated could fit spatially into MAL catalytic pocket, although they probably are weak or very transient binders and we identified molecular determinants involved in the binding of the substrate. The hydrophobic interactions formed by the methyl group of 3-methylaspartic acid, together with the presence of the amino group on carbon 2, play an essential role in the appropriate binding of the substrate. The results showed that β-lysine is able to fit and bind in MAL catalytic pocket and can be potentially converted from inhibitor to substrate of MAL upon enzyme engineering. The characterization of the binding and inhibition properties of the substrates tested here provide the foundation for future and more extensive studies on engineering MAL that could lead to a MAL variant able to catalyse this challenging deamination reaction.

Serum metabolite profiles as potential biochemical markers in young adults with community-acquired pneumonia cured by moxifloxacin therapy

Despite the utilization of various biochemical markers and probability calculation algorithms based on clinical studies of community-acquired pneumonia (CAP), more specific and practical biochemical markers remain to be found for improved diagnosis and prognosis. In this study, we aimed to detect the alteration of metabolite profiles, explore the correlation between serum metabolites and inflammatory markers, and seek potential biomarkers for young adults with CAP. 13 Eligible young mild CAP patients between the ages of 18 and 30 years old with CURB65 = 0 admitted to the respiratory medical department were enrolled, along with 36 healthy participants as control. Untargeted metabolomics profiling was performed and metabolites including alcohols, amino acids, carbohydrates, fatty acids, etc. were detected. A total of 227 serum metabolites were detected. L-Alanine, 2-Hydroxybutyric acid, Methylcysteine, L-Phenylalanine, Aminoadipic acid, L-Tryptophan, Rhamnose, Palmitoleic acid, Decanoylcarnitine, 2-Hydroxy-3-methylbutyric acid and Oxoglutaric acid were found to be significantly altered, which were enriched mainly in propanoate and tryptophan metabolism, as well as antibiotic-associated pathways. Aminoadipic acid was found to be significantly correlated with CRP levels and 2-Hydroxy-3-methylbutyric acid and Palmitoleic acid with PCT levels. The top 3 metabolites of diagnostic values are 2-Hydroxybutyric acid(AUC = 0.90), Methylcysteine(AUC = 0.85), and L-Alanine(AUC = 0.84). The AUC for CRP and PCT are 0.93 and 0.91 respectively. Altered metabolites were correlated with inflammation severity and were of great diagnostic value for CAP.

Untargeted metabolomics in halophytes: The role of different metabolites in New Zealand mangroves under multi-factorial abiotic stress conditions

Mangroves are halophytes which live in harsh coastal environments, and can serve as excellent model organisms to understand mechanisms of stress tolerance. Metabolomics is a useful approach to investigate the role different metabolites play during physiological responses of plants to abiotic stress factors. Previous studies indicate that levels of non-structural carbohydrates in plants may be associated with stress tolerance. Here, we manipulated levels of non-structural carbohydrates (NSCs) in the New Zealand mangrove, Avicennia marina subsp. australasica, through a light swapping regime. In a subsequent drought × salinity experiment, we then monitored leaf metabolite profiles of two NSC groups (high-NSC and low-NSC). Our results show that fourteen metabolites, belonging to multiple biochemical pathways, were significantly affected by one or more of these factors. The manipulation of non-structural carbohydrates led to increased abundance of amino acids in the low-NSC (L-NSC) plants compared to the high-NSC (H-NSC) plants. Under high drought conditions, the L-NSC plants had higher abundances of leucine and valine in comparison to those with the H-NSC phenotype. The L-NSC plants also had higher abundances of putrescine and aminoadipic acid when exposed to high salinity. Under the combination of drought and salinity, α-ketoglutarate was reduced in plants with the H-NSC phenotype, and soluble sugars accumulated compared to those with the L-NSC phenotype. The increased soluble sugar content in the H-NSC plants can facilitate osmotic adjustment, thereby aiding their survival during low soil water potential conditions. We also detected the presence of stress-protective phenolic compounds (syringic and sinapic acids) not previously reported in mangroves. Relationships between plant growth and the mangrove metabolome were also established, with levels of glucose and phosphoenol pyruvate being primarily responsible for phenotypic associations. Our findings show that the metabolites detected in this study, and their respective metabolic pathways, play a significant role in salinity and drought stress tolerance of A. marina subsp. australasica, providing new information to better understand mangrove mortality.