RESUMO
Considering the structure of the bacterial GH15 family glucoamylase (GA), Thermoplasma trehalase Tvn1315 may be composed of a ß-sandwich domain (BD) and a catalytic domain (CD). Tvn1315 BD weakly binds to insoluble ß-glucans, such as cellulose, and helps fold CD. To determine how aromatic residues contribute to proper folding and enzyme activity, we performed alanine scanning for 32 aromatic residues in the BD. The study did not identify a single residue involved in glucan binding. However, several aromatic residues were found to be involved in BD or CD folding and in modulating the activity of the full-length enzyme. Among those aromatic residue mutations, the W43A mutation led to reduced solubility of the BD and full-length protein and resulted in a full-length enzyme with significantly lower activity. The activity of W43F and W43Y was significantly higher than that of W43A. In addition, Ala substitutions of Tyr83, Tyr113, and Tyr17 led to a reduction in trehalase activity, but Phe substitutions of these residues could be tolerated, as these mutants maintained activities similar to WT activity. Thus, these aromatic residues in BD may interact with CD and modulate enzyme activity. KEY POINTS: ⢠Aromatic residues in the BD are involved in BD and CD folding. ⢠Aromatic residues in the BD near the CD active site modulate enzyme activity. ⢠BD interacts with CD and closely modulates enzyme activity.
Assuntos
Domínio Catalítico , Dobramento de Proteína , Trealase , Trealase/genética , Trealase/metabolismo , Trealase/química , Aminoácidos Aromáticos/metabolismo , Substituição de AminoácidosRESUMO
An asymmetric synthesis is a favorable approach for obtaining enantiomerically pure substances, but racemic resolution remains an efficient strategy. This study aims to elucidate the chiral resolution of aromatic amino acids and their elution order using glycopeptides as chiral selectors through molecular docking analysis. Chiral separation experiments were conducted using Vancomycin as a chiral additive in the mobile phase (CMPA) at various concentrations, coupled with an achiral amino column as the stationary phase. The Autodock Vina 1.1.2 software was employed to perform molecular docking simulations between each enantiomer (ligand) and Vancomycin (receptor) to evaluate binding affinities, demonstrate enantiomeric resolution feasibility, and elucidate chiral recognition mechanisms. Utilizing Vancomycin as CMPA at a concentration of 1.5 mM enabled the separation of tryptophan enantiomers with a resolution of 3.98 and tyrosine enantiomers with a resolution of 2.97. However, a poor chiral resolution was observed for phenylalanine and phenylglycine. Molecular docking analysis was employed to elucidate the lack of separation and elution order for tryptophan and tyrosine enantiomers. By calculating the binding energy, docking results were found to be in good agreement with experimental findings, providing insights into the underlying mechanisms governing chiral recognition in this system and the interaction sites. This comprehensive approach clarifies the complex relationship between chiral discrimination and molecular architecture, offering valuable information for creating and improving chiral separation protocols.
Assuntos
Aminoácidos Aromáticos , Glicopeptídeos , Simulação de Acoplamento Molecular , Glicopeptídeos/química , Aminoácidos Aromáticos/química , Estereoisomerismo , Vancomicina/química , Cromatografia Líquida de Alta Pressão/métodos , LigantesRESUMO
(1) Background: Branched-chain and aromatic amino acids (BCAAs/AAAs) have been considered as markers of type 2 diabetes (T2D); however, studies on associations between these metabolites and T2D and cardiometabolic traits in Hispanic populations are limited. The aim of this study was to examine the associations between baseline BCAAs (isoleucine, leucine, valine)/AAAs (phenylalanine, tyrosine) and prevalent and incident T2D, as well as baseline and longitudinal (2 year) changes in cardiometabolic traits (measures of glycemia, dyslipidemia, inflammation, and obesity) in two large cohorts of adults of Puerto Rican descent. (2) Methods: We included participants of the Boston Puerto Rican Health Study (BPRHS, n = 670) and San Juan Overweight Adult Longitudinal study (SOALS, n = 999) with available baseline metabolite and covariate data. T2D diagnosis was defined based on American Diabetes Association criteria. Multivariable logistic (for baseline T2D), Poisson (for incident T2D), and linear (for cardiometabolic traits) regression models were used; cohort-specific results were combined in the meta-analysis and adjusted for multiple comparisons. (3) Results: Higher baseline BCAAs were associated with higher odds of prevalent T2D (OR1SD BCAA score = 1.46, 95% CI: 1.34-1.59, p < 0.0001) and higher risk of incident T2D (IRR1SD BCAA score = 1.24, 95% CI: 1.13-1.37, p < 0.0001). In multivariable longitudinal analysis, higher leucine and valine concentrations were associated with 2-year increase in insulin (beta 1SD leucine = 0.37 mcU/mL, 95% CI: 0.11-0.63, p < 0.05; beta 1SD valine = 0.43 mcU/mL, 95% CI: 0.17-0.68, p < 0.01). Tyrosine was a significant predictor of incident T2D (IRR = 1.31, 95% CI: 1.09-1.58, p < 0.05), as well as 2 year increases in HOMA-IR (beta 1SD tyrosine = 0.13, 95% CI: 0.04-0.22, p < 0.05) and insulin concentrations (beta 1SD tyrosine = 0.37 mcU/mL, 95% CI: 0.12-0.61, p < 0.05). (4) Conclusions: Our results confirmed the associations between BCAAs and prevalent and incident T2D, as well as concurrent measures of glycemia, dyslipidemia, and obesity, previously reported in predominantly White and Asian populations. Baseline leucine, valine, and tyrosine were predictors of 2 year increases in insulin, whereas tyrosine was a significant predictor of deteriorating insulin resistance over time. Our study suggests that BCAAs and tyrosine could serve as early markers of future glycemic changes in Puerto Ricans.
Assuntos
Aminoácidos Aromáticos , Aminoácidos de Cadeia Ramificada , Fatores de Risco Cardiometabólico , Diabetes Mellitus Tipo 2 , Hispânico ou Latino , Humanos , Diabetes Mellitus Tipo 2/epidemiologia , Diabetes Mellitus Tipo 2/sangue , Feminino , Masculino , Pessoa de Meia-Idade , Aminoácidos de Cadeia Ramificada/sangue , Aminoácidos Aromáticos/sangue , Adulto , Hispânico ou Latino/estatística & dados numéricos , Estudos Longitudinais , Porto Rico/epidemiologia , Porto Rico/etnologia , Idoso , Prevalência , Boston/epidemiologia , Incidência , Obesidade/epidemiologia , Obesidade/etnologiaRESUMO
Although hydrophobic interactions provide the main driving force for initial peptide aggregation, their role in regulating suprastructure handedness of higher-order architectures remains largely unknown. We here interrogate the effects of hydrophobic amino acids on handedness at various assembly stages of peptide amphiphiles. Our studies reveal that relative to aliphatic side chains, aromatic side chains set the twisting directions of single ß-strands due to their strong steric repulsion to the backbone, and upon packing into multi-stranded ß-sheets, the side-chain aromatic interactions between strands form the aromatic ladders with a directional preference. This ordering not only leads to parallel ß-sheet arrangements but also induces the chiral flipping over of single ß-strands within a ß-sheet. In contrast, the lack of orientational hydrophobic interactions in the assembly of aliphatic peptides implies no chiral inversion upon packing into ß-sheets. This study opens an avenue to harness peptide aggregates with targeted handedness via aromatic side-chain interactions.
Assuntos
Interações Hidrofóbicas e Hidrofílicas , Peptídeos , Peptídeos/química , Peptídeos/metabolismo , Conformação Proteica em Folha beta , Estereoisomerismo , Estrutura Secundária de Proteína , Aminoácidos Aromáticos/química , Dicroísmo Circular , Modelos Moleculares , Aminoácidos/químicaRESUMO
Currently, urinary tract infection (UTI) diagnosis focuses on planktonic cell detection rather than biofilm detection, but the facile identification of UPEC bacterial biofilms is crucial in UTI diagnosis as the biofilm formed by bacteria is the causative agent of recurrent and chronic UTIs. Therefore, in this work, we developed a simple, cost-effective, colorimetric, and electrochemical-based strategy for the detection of cellulose in urine. Cellulose, a biofilm matrix component, was detected using tyrosine-capped gold and silver nanoparticles through a visible colorimetric change with a decrease in the absorbance intensity and a decrease in current response in the case of cyclic voltammetry. The sensor displayed a linear detection range of 10-70 mg mL-1 for colorimetry and 10-300 µg mL-1 for cyclic voltammetry with a good selectivity of <2.8% and a recovery rate of 95-100% in real-time sample analysis. Moreover, the binding affinity of cellulose with tyrosine was investigated using molecular docking studies to validate the sensing mechanism. We anticipate that our work will aid clinicians in the implementation of rapid, cost-effective, and definitive diagnosis of UTIs.
Assuntos
Biofilmes , Celulose , Ouro , Nanopartículas Metálicas , Prata , Infecções Urinárias , Infecções Urinárias/diagnóstico , Infecções Urinárias/microbiologia , Nanopartículas Metálicas/química , Prata/química , Ouro/química , Celulose/química , Humanos , Colorimetria/métodos , Diagnóstico Precoce , Aminoácidos Aromáticos/química , Técnicas Eletroquímicas , Simulação de Acoplamento MolecularRESUMO
Dimethylallyl tryptophan synthases (DMATSs) are aromatic prenyltransferases that catalyze the transfer of a prenyl moiety from a donor to an aromatic acceptor during the biosynthesis of microbial secondary metabolites. Due to their broad substrate scope, DMATSs are anticipated as biotechnological tools for producing bioactive prenylated aromatic compounds. Our study explored the substrate scope and product profile of a recombinant RePT, a novel DMATS from the thermophilic fungus Rasamsonia emersonii. Among a variety of aromatic substrates, RePT showed the highest substrate conversion for L-tryptophan and L-tyrosine (> 90%), yielding two mono-prenylated products in both cases. Nine phenolics from diverse phenolic subclasses were notably converted (> 10%), of which the stilbenes oxyresveratrol, piceatannol, pinostilbene, and resveratrol were the best acceptors (37-55% conversion). The position of prenylation was determined using NMR spectroscopy or annotated using MS2 fragmentation patterns, demonstrating that RePT mainly catalyzed mono-O-prenylation on the hydroxylated aromatic substrates. On L-tryptophan, a non-hydroxylated substrate, it preferentially catalyzed C7 prenylation with reverse N1 prenylation as a secondary reaction. Moreover, RePT also possessed substrate-dependent organic solvent tolerance in the presence of 20% (v/v) methanol or DMSO, where a significant conversion (> 90%) was maintained. Our study demonstrates the potential of RePT as a biocatalyst for the production of bioactive prenylated aromatic amino acids, stilbenes, and various phenolic compounds. KEY POINTS: ⢠RePT catalyzes prenylation of diverse aromatic substrates. ⢠RePT enables O-prenylation of phenolics, especially stilbenes. ⢠The novel RePT remains active in 20% methanol or DMSO.
Assuntos
Aminoácidos Aromáticos , Dimetilaliltranstransferase , Fenóis , Prenilação , Aminoácidos Aromáticos/metabolismo , Dimetilaliltranstransferase/metabolismo , Dimetilaliltranstransferase/genética , Fenóis/metabolismo , Especificidade por Substrato , Estilbenos/metabolismo , Triptofano/metabolismo , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/genéticaRESUMO
Aromatic amino acids (AAA) and derived compounds have enormous commercial value with extensive applications in the food, chemical and pharmaceutical fields. Microbial production of AAA and derived compounds is a promising prospect for its environmental friendliness and sustainability. However, low yield and production efficiency remain major challenges for realizing industrial production. With the advancement of synthetic biology, microbial production of AAA and derived compounds has been significantly facilitated. In this review, a comprehensive overview on the current progresses, challenges and corresponding solutions for AAA and derived compounds biosynthesis is provided. The most cutting-edge developments of synthetic biology technology in AAA and derived compounds biosynthesis, including CRISPR-based system, genetically encoded biosensors and synthetic genetic circuits, were highlighted. Finally, future prospects of modern strategies conducive to the biosynthesis of AAA and derived compounds are discussed. This review offers guidance on constructing microbial cell factory for aromatic compound using synthetic biology technology.
Assuntos
Aminoácidos Aromáticos , Biologia Sintética , Biologia Sintética/métodos , Aminoácidos Aromáticos/biossíntese , Engenharia Metabólica/métodos , Técnicas Biossensoriais/métodos , Bactérias/metabolismo , Bactérias/genéticaRESUMO
Amphiphilic peptides show great potential for exfoliating graphite and functionalizing graphene. However, the variety of amino acids complicates our understanding of the underlying mechanisms. In this study, we designed four peptides (C6W1, C6W2, C6W4, and C6W6) with different amounts of aromatic tryptophan amino acids and two additional peptides (C6F4 and C6Y4) by substituting tryptophan with aromatic phenylalanine or tyrosine. This allowed us to investigate the processes and mechanisms of graphite exfoliation and graphene functionalization. Using experimental and computational methods, we discovered that peptides containing tryptophan demonstrated higher exfoliation efficiency and increased tryptophan content further improved this efficiency, resulting in more peptide-functionalized graphene layers. Significantly, the primary driving force for the surface-assisted assembly of peptides on graphite is the π-π stacking interaction between the aromatic ring contributed by aromatic amino acids and the hexagonal rings of the graphite surface. This interaction leads to a layer-by-layer exfoliation mechanism. Our research offers valuable insights into peptide design strategies for one-step graphite exfoliation and graphene functionalization in aqueous environments.
Assuntos
Aminoácidos Aromáticos , Grafite , Peptídeos , Propriedades de Superfície , Grafite/química , Peptídeos/química , Aminoácidos Aromáticos/química , Triptofano/química , Tensoativos/químicaRESUMO
Aromatic d-amino acids (d-AAs) play a pivotal role as important chiral building blocks and key intermediates in fine chemical and drug synthesis. Meso-diaminopimelate dehydrogenase (DAPDH) serves as an excellent biocatalyst in the synthesis of d-AAs and their derivatives. However, its strict substrate specificity and the lack of efficient engineering methods have hindered its widespread application. Therefore, this study aims to elucidate the catalytic mechanism underlying DAPDH from Proteus vulgaris (PvDAPDH) through the examination of its crystallographic structure, computational simulations of potential energies and molecular dynamics simulations, and site-directed mutagenesis. Mechanism-guided computational design showed that the optimal mutant PvDAPDH-M3 increased specific activity and catalytic efficiency (kcat/Km) for aromatic keto acids up to 124-fold and 92.4-fold, respectively, compared to that of the wild type. Additionally, it expanded the substrate scope to 10 aromatic keto acid substrates. Finally, six high-value-added aromatic d-AAs and their derivatives were synthesized using a one-pot three-enzyme cascade reaction, exhibiting a good conversion rate ranging from 32 to 84% and excellent stereoselectivity (enantiomeric excess >99%). These findings provide a potential synthetic pathway for the green industrial production of aromatic d-AAs.
Assuntos
Aminoácido Oxirredutases , Aminoácidos Aromáticos , Simulação de Dinâmica Molecular , Mutagênese Sítio-Dirigida , Aminoácido Oxirredutases/metabolismo , Aminoácido Oxirredutases/genética , Aminoácido Oxirredutases/química , Especificidade por Substrato , Aminoácidos Aromáticos/metabolismo , Aminoácidos Aromáticos/biossíntese , Proteus vulgaris/enzimologia , Proteus vulgaris/genética , Biocatálise , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/químicaRESUMO
Ingestion of fermented foods impacts human immune function, yet the bioactive food components underlying these effects are not understood. Here, we interrogated whether fermented food bioactivity relates to microbial metabolites derived from aromatic amino acids, termed aryl-lactates. Using targeted metabolomics, we established the presence of aryl-lactates in commercially available fermented foods. After pinpointing fermented food-associated lactic acid bacteria that produce high levels of aryl-lactates, we identified fermentation conditions to increase aryl-lactate production in food matrices up to 5 × 103 fold vs. standard fermentation conditions. Using ex vivo reporter assays, we found that food matrix conditions optimized for aryl-lactate production exhibited enhanced agonist activity for the human aryl-hydrocarbon receptor (AhR) as compared to standard fermentation conditions and commercial products. Reduced microbial-induced AhR activity has emerged as a hallmark of many chronic inflammatory diseases, thus we envision strategies to enhance AhR bioactivity of fermented foods to be leveraged to improve human health.
Assuntos
Aminoácidos Aromáticos , Fermentação , Alimentos Fermentados , Receptores de Hidrocarboneto Arílico , Humanos , Alimentos Fermentados/análise , Alimentos Fermentados/microbiologia , Aminoácidos Aromáticos/metabolismo , Receptores de Hidrocarboneto Arílico/metabolismo , Lactobacillales/metabolismo , Lactatos/metabolismoRESUMO
In the face of ongoing water pollution challenges, the intricate interplay between dissolved organic matter and disinfectants like chlorine gives rise to potentially harmful disinfection byproducts (DBPs) during water treatment. The exploration of DBP formation originating from amino acids (AA) is a critical focus of global research. Aromatic DBPs, in particular, have garnered considerable attention due to their markedly higher toxicity compared to their aliphatic counterparts. This work seeks to advance the understanding of DBP formation by investigating chlorination disinfection and kinetics using tyrosine (Tyr), phenylalanine (Phe), and tryptophan (Trp) as precursors. Via rigorous experiments, a total of 15 distinct DBPs with accurate molecular structures were successfully identified. The chlorination of all three AAs yielded highly toxic chlorophenylacetonitriles (CPANs), and the disinfectant dosage and pH value of the reaction system potentially influence chlorination kinetics. Notably, Phe exhibited the highest degradation rate compared to Tyr and Trp, at both the CAA:CHOCl ratio of within 1:2 and a wide pH range (6.0 to 9.0). Additionally, a neutral pH environment triggered the maximal reaction rates of the three AAs, while an acidic condition may reduce their reactivity. Overall, this study aims to augment the DBP database and foster a deeper comprehension of the DBP formation and relevant kinetics underlying the chlorination of aromatic AAs.
Assuntos
Aminoácidos Aromáticos , Desinfecção , Halogenação , Purificação da Água , Cinética , Aminoácidos Aromáticos/química , Purificação da Água/métodos , Desinfetantes/química , Poluentes Químicos da Água/química , Concentração de Íons de HidrogênioRESUMO
BACKGROUND: Malnutrition is a common geriatric syndrome that is closely associated with adverse clinical outcomes and poses significant harm to older adults. Early assessment of nutritional status plays a crucial role in preventing and intervening in cases of malnutrition. However, there is currently a lack of measurable methods and biomarkers to evaluate malnutrition in older adults accurately. The aim of this study is to investigate the independent correlation between serum levels of amino acids and malnutrition in older adults, and to identify effective metabolomics biomarkers that can aid in the early detection of geriatric malnutrition. METHODS: A total of 254 geriatric medical examination participants from Beijing Hospital were included in the study, consisting of 182 individuals with normal nutritional status (Normal group) and 72 patients at risk of malnutrition or already malnourished (MN group). Malnutrition was assessed using the Mini-Nutritional Assessment Short-Form (MNA-SF). Demographic data were collected, and muscle-related and lipid indexes were determined. Serum amino acid concentrations were measured using isotope dilution liquid chromatography-tandem mass spectrometry (LC-MS/MS). The correlation between serum amino acid levels and malnutrition was analyzed using non-parametric tests, partial correlation analysis, linear regression, and logistic regression. RESULTS: The geriatric MN group exhibited significantly lower serum aromatic amino acid levels (P < 0.05) compared to the normal group. A positive correlation was observed between serum aromatic amino acid levels and the MNA-SF score (P = 0.002), as well as with known biomarkers of malnutrition such as body mass index (BMI) (P < 0.001) and hemoglobin (HGB) (P = 0.005). Multivariable logistic or linear regression analyses showed that aromatic amino acid levels were negatively correlated with MN and positively correlated with the MNA-SF score, after adjusting for some confounding factors, such as age, gender, BMI, smoking status, history of dyslipidemia, diabetes mellitus and frailty. Stratified analyses revealed that these trends were more pronounced in individuals without a history of frailty compared to those with a history of frailty, and there was an interaction between aromatic amino acid levels and frailty history (P = 0.004). CONCLUSION: Our study suggests that serum aromatic amino acids are independently associated with malnutrition in older adults. These results have important implications for identifying potential biomarkers to predict geriatric malnutrition or monitor its progression and severity, as malnutrition can result in poor clinical outcomes.
Assuntos
Fragilidade , Desnutrição , Humanos , Idoso , Fragilidade/diagnóstico , Cromatografia Líquida , Espectrometria de Massas em Tandem , Desnutrição/diagnóstico , Desnutrição/complicações , Estado Nutricional , Avaliação Nutricional , Biomarcadores , Aminoácidos , Aminoácidos Aromáticos , Avaliação Geriátrica/métodosRESUMO
Nitrogen-doped carbon dots (NCD) were synthesized using a simple and fast hydrothermal route, employing citric acid and urea as precursors. The resulting NCDs were non-covalently functionalized (conjugated) with aromatic amino acids, namely phenylalanine (Phe) and tryptophan (Trp). Atomic force microscopy revealed that the NCDs exhibit a disk-like morphology with an average diameter of approximately 60â¯nm and an average height of about 0.5â¯nm. Following conjugation, the particle height increased to around 3â¯nm. UV-vis spectroscopy analysis indicated successful conjugation of the amino acids to the NCD nanostructures. Additionally, DFT numerical calculations based on three differently N-doped clusters were performed to elucidate the nature of the non-covalent interactions between NCDs and the corresponding amino acids. Photoluminescent spectra demonstrated a stable and strong fluorescence signal for both hybrids in the UV region. The most significant changes were observed in the case of Trp-conjugation. In contrast to phenylalanine, the non-covalent bonding of tryptophan to NCDs strongly influenced the visible emission (around 500â¯nm) originating from surface states of the dots.
Assuntos
Aminoácidos Aromáticos , Carbono , Nanoestruturas , Nitrogênio , Carbono/química , Nitrogênio/química , Aminoácidos Aromáticos/química , Nanoestruturas/química , Pontos Quânticos/química , Propriedades de Superfície , Fenilalanina/química , Tamanho da Partícula , Triptofano/química , Microscopia de Força Atômica , Fenômenos Ópticos , Teoria da Densidade FuncionalRESUMO
Protein phosphorylation is a prevalent translational modification, and its dysregulation has been implicated in various diseases, including cancer. Despite its significance, there is a lack of specific inhibitors of the FCP/SCP-type Ser/Thr protein phosphatase Scp1, characterized by high specificity and affinity. In this study, we focused on adnectin, an antibody-mimetic protein, aiming to identify Scp1-specific binding molecules with a broad binding surface that target the substrate-recognition site of Scp1. Biopanning of Scp1 was performed using an adnectin-presenting phage library with a randomized FG loop. We succeeded in identifying FG-1Adn, which showed high affinity and specificity for Scp1. Ala scanning analysis of the Scp1-binding sequence in relation to the FG-1 peptide revealed that hydrophobic residues, including aromatic amino acids, play important roles in Scp1 recognition. Furthermore, FG-1Adn was found to co-localize with Scp1 in cells, especially on the plasma membrane. In addition, Western blotting analysis showed that FG-1Adn increased the phosphorylation level of the target protein of Scp1 in cells, indicating that FG-1Adn can inhibit the function of Scp1. These results suggest that FG-1Adn can be used as a specific inhibitor of Scp1.
Assuntos
Anticorpos , Domínio de Fibronectina Tipo III , Proteínas Recombinantes , Aminoácidos Aromáticos , Fosfoproteínas Fosfatases , Biblioteca de PeptídeosRESUMO
BACKGROUND: Obesity is a progressive metabolic disease that begins with lipid metabolism disorders. Aromatic amino acids (AAAs), including tryptophan, phenylalanine, and tyrosine, have diverse biological activities as nutrients. However, the underlying mechanisms by which AAAs affect lipid metabolism are unclear. OBJECTIVES: This study was designed to investigate the possible roles and underlying molecular mechanisms of AAA in the pathogenesis of lipid metabolism disorders. METHODS: We added an AAA mixture to the high-fat diet (HFD) of mice. Glucose tolerance test was recorded. Protein expression of hepatic bile acid (BA) synthase and mRNA expression of BA metabolism-related genes were determined. Hepatic BA profiles and gut microbial were also determined in mice. RESULTS: The results showed that AAA significantly increased body weight and white adipose tissue, aggravated liver injury, impaired glucose tolerance and intestinal integrity, and significantly increased hepatic BA synthesis by inhibiting intestinal farnesoid X receptor (FXR). Moreover, AAA increased the content of total BA in the liver and altered the hepatic BA profile, with elevated levels of lithocholic acid, glycochenodeoxycholic acid, and glycoursodeoxycholic acid. AAA markedly increased the levels of proteins involved in BA synthesis (cholesterol 7α-hydroxylase and oxysterol 7α-hydroxylase) and inhibited the intestinal FXR. Gut microbial composition also changed, reducing the abundance of some beneficial bacteria, such as Parvibacter and Lactobacillus. CONCLUSIONS: Under HFD conditions, AAAs stimulate BA synthesis in both the classical and alternative pathways, leading to aggravation of liver injury and fat deposition. Excessive intake of AAA disrupts BA metabolism and contributes to the development of lipid metabolism disorders, suggesting that AAA may be a causative agent of lipid metabolism disorders.
Assuntos
Transtornos do Metabolismo dos Lipídeos , Metabolismo dos Lipídeos , Camundongos , Animais , Aminoácidos Aromáticos , Fígado/metabolismo , Transtornos do Metabolismo dos Lipídeos/metabolismo , Ácidos e Sais Biliares/metabolismo , Camundongos Endogâmicos C57BLRESUMO
In this study, we investigated the trimerization mechanism and structure of heat shock factor 1 (HSF1) using western blotting, tryptophan (Trp) fluorescence spectroscopy, and molecular modeling. First, we examined the DNA-binding domains of human (Homo sapiens), goldfish (Carassius auratus), and walleye pollock (Gadus chalcogrammus) HSF1s by mutating key residues (36 and 103) that are thought to directly affect trimer formation. Human, goldfish, and walleye pollock HSF1s contain cysteine at residue 36 but cysteine (C), tyrosine (Y), and phenylalanine (F), respectively, at residue 103. The optimal trimerization temperatures for the wild-type HSF1s of each species were found to be 42, 37, and 20 °C, respectively. Interestingly, a mutation experiment revealed that trimerization occurred at 42 °C when residue 103 was cysteine, at 37 °C when it was tyrosine, and at 20 °C when it was phenylalanine, regardless of the species. In addition, it was confirmed that when residue 103 of the three species was mutated to alanine, trimerization did not occur. This suggests that in addition to trimerization via disulfide bond formation between the cysteine residues in human HSF1, trimerization can also occur via the formation of a different type of bond between cysteine and aromatic ring residues such as tyrosine and phenylalanine. We also confirmed that at least one cysteine is required for the trimerization of HSF1s, regardless of its position (residue 36 or 103). Additionally, it was shown that the trimer formation temperature is related to growth and survival in fish.
Assuntos
Aminoácidos Aromáticos , Cisteína , Fatores de Transcrição de Choque Térmico , Animais , Humanos , Aminoácidos Aromáticos/metabolismo , Aminoácidos Aromáticos/química , Cisteína/química , Cisteína/metabolismo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Carpa Dourada/metabolismo , Fatores de Transcrição de Choque Térmico/metabolismo , Fatores de Transcrição de Choque Térmico/química , Fatores de Transcrição de Choque Térmico/genética , Modelos Moleculares , Domínios Proteicos , Multimerização ProteicaRESUMO
Octanal was found to be able to reduce green mold incidence in citrus fruit by a defense response mechanism. However, the underlying mechanism remains largely unclear. Herein, the metabolomics, RNA-seq and biochemical analyses were integrated to explore the effect of octanal on disease resistance in harvested citrus fruit. Results showed that octanal fumigation at 40 µL L-1 was effective in controlling citrus green mold. Metabolomics analysis showed that octanal mainly led to the accumulation of some plant hormones including methyl jasmonate, abscisic acid, indole-3-butyric acid, indoleacetic acid (IAA), salicylic acid, and gibberellic acid and many phenylpropanoid metabolites including cinnamyl alcohol, hesperidin, dihydrokaempferol, vanillin, quercetin-3-O-malonylglucoside, curcumin, naringin, chrysin, coniferin, calycosin-7-O-ß-D-glucoside, trans-cinnamaldehyde, and 4',5,7-trihydroxy-3,6-dimethoxyflavone. Particularly, IAA and hesperidin were dramatically accumulated in the peel, which might be the contributors to the resistance response. Additionally, transcriptome analysis showed that octanal greatly activated the biosynthesis and metabolism of aromatic amino acids. This was further verified by the accumulation of some metabolites (shikimic acid, tryptophan, tyrosine, phenylalanine, IAA, total phenolics, flavonoids and lignin), increase in some enzyme activities (phenylalanine ammonia-lyase, tyrosine ammonia-lyase, 4-coumarate CoA ligase, cinnamic acid 4-hydroxylase, polyphenol oxidase, and peroxidase), up-regulation of some genes (tryptophan pyruvate aminotransferase, aldehyde dehydrogenase, shikimate kinase and shikimate dehydrogenase) expressions and molecular docking results. Thus, these results indicate that octanal is an efficient strategy for the control of postharvest green mold by triggering the defense response in citrus fruit.
Assuntos
Aldeídos , Citrus , Hesperidina , Citrus/química , Citrus/genética , Citrus/metabolismo , Aminoácidos Aromáticos/metabolismo , Resistência à Doença , Hesperidina/análise , Hesperidina/metabolismo , Hesperidina/farmacologia , Triptofano/metabolismo , Simulação de Acoplamento Molecular , FrutasRESUMO
The proteasome is a multisubunit protease system responsible for the majority of the protein turnover in eukaryotic organisms. Dysregulation of this enzymatic complex leads to protein accumulation, subsequent aggregation, and ultimately diseased states; for that reason, positive modulation of its activity has been recently investigated as a therapeutic strategy for neurodegenerative and age-related diseases. The small molecule AM404 was recently identified as an activator of the 20S isoform of the proteasome and further exploration of the scaffold revealed the importance of the polyunsaturated fatty acid chain to elicit activity. Herein, we report the investigation of the aromatic region of the scaffold and the evaluation of the small molecules in a variety of proteasome activity and protein degradation assays. We found that derivatives A22 and A23, compared to AM404, exhibit enhanced proteasome activity in biochemical and cellular proteasome assays and more favorable cellular viability profiles. Additionally, these compounds demonstrate the ability to degrade intrinsically disordered proteins, regardless of their molecular weight, and the ability to restore the proteasome activity in the presence of toxic oligomeric α-Syn species in a biochemical setting.
Assuntos
Ácidos Araquidônicos , Ativadores de Enzimas , Complexo de Endopeptidases do Proteassoma , Ácidos Araquidônicos/química , Ácidos Araquidônicos/farmacologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Ativadores de Enzimas/metabolismo , Ativadores de Enzimas/farmacologia , Ativação Enzimática/efeitos dos fármacos , Proteólise/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Proteínas Intrinsicamente Desordenadas/metabolismo , Aminoácidos Aromáticos/metabolismoRESUMO
Aromatic amino acids (AAAs) are essential for synthesis of proteins and numerous plant natural products, yet how plants maintain AAA homeostasis remains poorly understood. Wu et al. reported that the aminotransferase VAS1 plays a role in AAA homeostasis by transferring nitrogen from AAAs to non-proteinogenic amino acids, 3-carboxytyrosine and 3-carboxyphenylalanine.
Assuntos
Aminoácidos Aromáticos , Homeostase , Nitrogênio , Aminoácidos Aromáticos/metabolismo , Nitrogênio/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Transaminases/metabolismoRESUMO
The aromatic amino acids tryptophan, phenylalanine, and tyrosine are targets for oxidation during food processing. We investigated whether S. cerevisiae can use nonproteinogenic aromatic amino acids as substrates for degradation via the Ehrlich pathway. The metabolic fate of seven amino acids (p-, o-, m-tyrosine, 3,4-dihydroxyphenylalanine (DOPA), 3-nitrotyrosine, 3-chlorotyrosine, and dityrosine) in the presence of S. cerevisiae was assessed. All investigated amino acids except dityrosine were metabolized by yeast. The amino acids 3-nitrotyrosine and o-tyrosine were removed from the medium as fast as p-tyrosine, and m-tyrosine, 3-chlorotyrosine, and DOPA more slowly. In summary, 11 metabolites were identified by high-performance liquid chromatography-mass spectrometry (HPLC-MS/MS). DOPA, 3-nitrotyrosine, and p-tyrosine were metabolized predominantly to the Ehrlich alcohols, whereas o-tyrosine and m-tyrosine were metabolized predominantly to α-hydroxy acids. Our results indicate that nonproteinogenic aromatic amino acids can be taken up and transaminated by S. cerevisiae quite effectively but that decarboxylation and reduction to Ehrlich alcohols as the final metabolites is hampered by hydroxyl groups in the o- or m-positions of the phenyl ring. The data on amino acid metabolism were substantiated by the analysis of five commercial beer samples, which revealed the presence of hydroxytyrosol (ca. 0.01-0.1 mg/L) in beer for the first time.