Among the 20 standard amino acids, aromatic amino acids hold particular importance due to their crucial roles in protein–protein interactions. These residues, including phenylalanine, tyrosine, tryptophan, and histidine, contribute significantly to protein structure and function through their ability to engage in non-covalent interactions, such as π–π stacking and cation–π interactions. In addition, the acid–base duality of aromatic amino acids makes their interactions with metal ions particularly valuable. These interactions are implicated in several essential biological processes, including protein biosynthesis, enzyme catalysis, and molecular recognition. While the interactions between transition metal ions and amino acids have been extensively studied and well-documented, the role of alkali metal ions in similar contexts has received comparatively less attention. However, their influence on amino acid coordination and stability cannot be overlooked, especially given their biological relevance in processes such as ionic regulation, enzyme activation, and membrane transport. The results of this study demonstrate that the complexation of aromatic amino acids with alkali metal ions enhances their acidity. Additionally, we examined the detailed interactions of these metals with the aromatic amino acids.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00726-025-03486-x.

L-arginine (L-Arg) is metabolised in the cell to generate nitric oxide (NO) and citrulline via nitric oxide synthase (NOS). NO is an important cellular signalling molecule that regulates lipid and glucose metabolism. The biological availability of NO is affected by the NOS inhibitor; NG-nitro-L-Arg methyl ester (L-NAME) and the external NO donor; S-nitroso-N-acetyl-D, L-penicillamine (SNAP). Mouse adipocyte 3T3 L1 cells were cultured with 0, 400 and 800 µM L-Arg or control complete DMEM media. The impact of L-NAME (4 mM), and SNAP (100 µM) was also analysed. The cell fitness was similar and the mRNA levels of AMPK was increased and ACC-1 was decreased, whilst the activation of AMPK and ACC-1 was decreased upon the addition of exogenous L-Arg. Transcript and protein levels of AMPK and ACC-1 were regulated by addition of L-NAME and SNAP, however the impact of these targets was related to the concentration of L-Arg added to the cells and the culture time point of analysis. NO in the form of NO2− in cell culture supernatant was elevated in 400 and 800 µM L-Arg cultures. L-NAME significantly inhibited NO production from adipose cells in a time-dependent manner and subsequently impacted AMPK and ACC expression. Associated with these changes were changed in the concentration of L-Arg, L-Cit and L-Orn in the culture media. Collectively, these results show that excess L-Arg is sensed by the cell which then regulates AMPK and ACC-1 expression in response. The findings could have implications in modulation of signalling pathways for treating obesity and obesity induced diabetic mellitus.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00726-025-03467-0.

l-threonine is used in dietary supplements and nutritional products ingested by healthy consumers. The objective of this study was to determine in a randomized double blind controlled clinical trial the safety and tolerability of l-threonine used as graded doses in supplements for 4 weeks. Healthy male adults (age 42.9) ingested randomly placebo or different doses of L-threonine (0, 3, 6, 9, 12 g/day) for 4 weeks using a crossover design. At the end of supplementation period, the subjects visited the clinic for medical examination, anthropometric parameter measurements, blood sampling for biochemical tests including amino acid concentrations in plasma, measurement of blood pressure and heart rate, and dietary intake evaluation. Adverse events were recorded all along the trial. None of the anthropometric parameters measured, dietary intake and the biochemical parameters were affected by l-threonine supplementation except a non-specific minor increase in plasma aspartate amino transferase and creatine kinase which was measured in the group supplemented with 9 g l-threonine per day but not with the 12 g per day dose. Also, the concentration of L-threonine as well as the concentration of its metabolite L-2-amino butylate were found to be increased in plasma after supplementation with 6, 9, 12 g/day L-threonine. The moderate and mild adverse events were found to occur at random. All symptoms disappeared during the supplementation period despite continuous L-threonine supplementation. These results of this study indicate a no-observed-adverse-effect-level (NOAEL) value for L-threonine to be 12 g/day in healthy adult males. This study was registered at jRCT as jRCT1050230137.

Compound-specific stable carbon isotope analysis of amino acids (CSIA-AA) is widely used in ecological studies to analyze food-webs and is gaining use in archaeology for investigating past diets. However, its use in reconstructing breastfeeding and weaning practices is not fully understood. This study evaluates the efficacy of stable carbon isotope analysis of amino acids in early life diet reconstruction by analyzing keratin from fingernail samples of three mother-infant pairs during late gestation and early postpartum periods. Our results show that stable carbon isotope ratios (δ13C) of glycine, and to a lesser extent glutamate, effectively trace the onset of exclusive breastfeeding and the end of weaning in infants. We propose that glycine’s ‘conditionally essential’ metabolic pathway during infancy allows it to reflect maternal glycine δ13C, indicating breastmilk consumption. Subtle changes in glutamate δ13C likely result from its ‘non-essential’ status. Additionally, δ13C values of glycine and glutamate indicate maternal physiological and pathological stress due to catabolic effects such as gluconeogenesis. These findings have significant implications for ecological and archaeological research using CSIA-AA for dietary reconstructions. They highlight the need to understand how metabolic pathways affecting δ13C of amino acids may change over an individual’s lifespan or be altered due to various forms of stress.

Four aliphatic amino acids—α-aminobutyric acid (AABA), β-aminobutyric acid (BABA), α-aminoisobutyric acid (AAIBA) and β-aminoisobutyric acid (BAIBA) were investigated in water as a solvent by two quantum chemical methods. B3LYP hybrid version of DFT was used for geometry optimization and a full vibrational analysis of neutral molecules, their cations and anions in the canonical and zwitterionic forms (6 forms for each species). Ab initio DLPNO-CCSD(T) method was applied in the geometry pre-optimized by B3LYP. Calculated molecular descriptors involve dipole moment, quadrupole moment, dipole polarizability, energy of zero-point vibration and total entropic term which enter the standard Gibbs energy. In addition, a set of collective electronic and thermodynamic properties associated with redox process were evaluated: ionization energy, electron affinity, chemical hardness, molecular electronegativity, electrophilicity index, absolute oxidation and reduction potentials. A mutual comparison of these structural isomers including γ-aminobutyric acid (GABA) shows high degree of similarity in molecular descriptors. However, cluster analysis of 12 electro neutral, linear and branched amino acids with 2 – 6 carbon atoms discriminates them into five clusters. It is found that the electrophilicity index correlates with the absolute reduction potential along a straight line (24 items). The reduction potential for canonical structure varies between 1.21 V (glycine) and 1.45 V (AABA) whereas for the zwitterionic form it is visibly lower 0.52–1.11 V. The highest absolute reduction potential > 1.43 V is shown by α-amino acids: α-alanine, AABA (homoalanine) and AAIBA having 2-methyl or 2-ethyl functional group. The calculated absolute oxidation potential correlates with the adiabatic ionization energy and can be used as a criterion of the antioxidant capacity. According to thermodynamic data, the SPLET mechanism of the electron-proton coupled transfer is favored over the alternative SET-PT mechanism. This work contributes to the creation of a database of molecular properties of amino acids based on the same method and basis set.

Plants are known as a source of different biologically active compounds, which are uncommon for other kingdoms of life. Among them are different amino acid analogues, which are synthesized and accumulated in certain plants as a passive defense mechanism against herbivorous insects and grazing mammals. As a rule, cell protein synthesis machinery of herbivores cannot effectively differentiate between standard proteinogenic amino acids and their specific plant analogues, resulting in misincorporation of the latter into nascent proteins and their malfunctioning, which constitutes a mechanism of plant defense. Examples of such amino acids are analogues of arginine (canavanine, indospicine), proline (azetidine-2-carboxylic acid), and cysteine/lysine (thialyasine). This review summarizes existing knowledge on these and other related amino acids as potential antibacterial and antifungal agents, including their possible targets and known resistance mechanisms. We also discuss the possibility of using amino acid analogues as sole antimicrobial agents or in combination with known antibacterials and antifungals. We also propose a strategy of enhancing the antimicrobial activity of amino acid analogue by concomitant starvation for the corresponding standard amino acid, which has been proven efficient in anticancer studies. Such an approach might potentially help to overcome, at least partially, microbial resistance to known antibiotics, especially when such resistance relies on increased protein synthesis in pathogen cells.

Claudin (CLDN) proteins are extensively studied due to their critical role in maintaining tissue barriers and cell polarity. However, significant gaps remain in understanding the functional mechanisms of their sequence motifs and the molecular mechanisms of their interactions with other tight junction proteins. This review systematically examines the multifunctional properties of the CLDN protein family from the perspectives of sequence and structure. During evolution, CLDN family members have developed highly conserved structural features, particularly key conserved sites within the first extracellular loop (ECL1) and the C-terminal PDZ-binding domain, which play a central role in regulating the barrier function of tight junctions, ion selectivity, and protein–protein interactions. Furthermore, the distribution pattern of acidic and basic amino acids in ECL1 has been shown to directly determine ion selectivity and paracellular permeability. Meanwhile, the assembly and functional stability of tight junctions are precisely regulated by the C-terminal PDZ-binding domain through its interactions with the ZO protein family. Additionally, the study further elucidates how CLDN proteins modulate critical signaling pathways governing cellular proliferation, survival, and permeability, thereby participating in diverse physiological and pathological processes. These insights have deepened the understanding of the functional diversity of CLDN proteins and provided a new theoretical basis for developing disease diagnostic markers and designing targeted treatment strategies based on CLDN proteins.

Designing innovative, accurate, universal, and accessible diagnostic tests is mandatory to improve screening, prevention, and management of hepatitis D (HD), especially in endemic areas with poor infrastructure and restricted access to public health care. Recombinant proteins (RP), recombinant multiepitope proteins (RMP), and synthetic peptides have been extensively reported as tools for efficient immunodiagnosis of several diseases. This review aimed to discuss the use of these antigens for the immunodiagnosis of HD. To this end, a bibliographic study was conducted in the PubMed database by searching the primary (“Hepatitis D” and “Hepatitis Delta”), secondary (“Detection”, “Diagno*”, “Diagnosis”, “Immunodiagnosis”, and “Serodiagnosis”), and tertiary (“Chimera”, “Epitope”, “Peptide”; “Protein” and “Recombinant”) descriptors, including papers published up to January 2025. Review articles and case reports were excluded. Only nine articles (five for RP, three for synthetic peptides, and one for RMP) met the inclusion criteria, revealing that there are very few studies on this subject, particularly when compared to the advances made in the diagnosis of hepatitis A, B, and C. Despite the scarcity of articles published in the literature, six of the nine analyzed studies corroborate the potential of these antigens to effectively replace traditional diagnostic methods, including development of rapid tests. These data highlight the need for further studies to assess the potential of RP, RMP, and synthetic peptides for immunodiagnosis of HD, aiming to increase the accuracy of diagnosis, as well as improve monitoring and prevention.

The obesity epidemic among children has become a major public health issue, and the presence of childhood insulin resistance (IR) has been demonstrated prior to the onset of type 2 diabetes mellitus. However, it is unclear whether the metabolomic signature is associated with weight loss interventions in obese children with IR. Thirty-six obese children with IR were selected from the weight loss camp (Shenzhen Sunshine Xing Yada health Technology Co., LTD). Clinical parameters were collected before and after weight loss intervention. Targeted metabolomics of plasma samples was performed by ultra-performance liquid chromatography coupled to the tandem mass spectrometry, and principal component analysis, variable importance in projection, and orthogonal partial least squares discriminant analysis were used to obtain the differentially expressed metabolites. Pathway analysis was conducted with the Homo sapiens (HSA) sets in the Kyoto Encyclopedia of Genes and Genomes. We used machine learning algorithms to obtain the potential biomarkers and Spearman correlation analysis to clarify the association between potential biomarkers and clinical parameters. We found that clinical parameters and metabolite clusters were significantly changed in obese children with IR before and after weight loss intervention. Mechanistically, weight loss intervention significantly changed 61 metabolites in obese children with IR. Furthermore, 12 pathways were significantly changed. Moreover, the machine learning algorithm found 6 important potential biomarkers. In addition, these potential biomarkers were strongly associated with major clinical parameters. These data indicate different metabolomic profiles in obese children with IR after weight loss intervention, providing insights into the clinical parameters and metabolite mechanisms involved in weight loss programs.

Studies in vivo have demonstrated that the accumulation of d-amino acids (d-AAs) is associated with age-related diseases and increased immune activation. However, the underlying mechanism(s) of these observations are not well defined. The metabolism of d-AAs by d-amino oxidase (DAO) produces hydrogen peroxide (H2O2), a reactive oxygen species involved in several physiological processes including immune response, cell differentiation, and proliferation. Excessive levels of H2O2 contribute to oxidative stress and eventual cell death, a characteristic of age-related pathology. Here, we explored the molecular mechanisms of d-serine (d-Ser) and d-alanine (d-Ala) in human liver cancer cells, HepG2, with a focus on the production of H2O2 the downstream secretion of pro-inflammatory cytokine and chemokine, and subsequent cell death. In HepG2 cells, we demonstrated that d-Ser decreased H2O2 production and induced concentration-dependent depolarization of mitochondrial membrane potential (MMP). This was associated with the upregulation of activated NF-кB, pro-inflammatory cytokine, TNF-α, and chemokine, IL-8 secretion, and subsequent apoptosis. Conversely, d-Ala-treated cells induced H2O2 production, and were also accompanied by the upregulation of activated NF-кB, TNF-α, and IL-8, but did not cause significant apoptosis. The present study confirms the role of both d-Ser and d-Ala in inducing inflammatory responses, but each via unique activation pathways. This response was associated with apoptotic cell death only with d-Ser. Further research is required to gain a better understanding of the mechanisms underlying d-AA-induced inflammation and its downstream consequences, especially in the context of aging given the wide detection of these entities in systemic circulation.