Key Amino Acid Research Articles

A Cryptic Loop that Modulates Protein-Protein Interaction: Implications in PfAMA1-PfRON2 Late-stage Binding Event.

Disease-related protein-protein interactions are important molecular targets for drug discovery campaigns. The dynamics of a disordered loop, which are commonly found in the receptor binding domain of many proteins, often plays a decisive role during protein-protein or protein-small molecule binding events. One notable example is the interactions between two proteins from Plasmodium falciparum, PfAMA1-PfRON2, which are crucial for the malaria parasite's invasion into human red blood cells. A thorough understanding of the interactions between these macromolecular binding partners is important for designing better therapeutics against this ancient disease. The available crystal structures of the PfAMA1-PfRON2 complex show insufficient electron density, making it challenging to fully understand the molecular-level association of the domain II (DII) loop with the PfRON2. To address this, we have computationally simulated the dynamics and free energetics of DII loop closing processes, identifying a set of key amino acid residues in the PfRON2 helix that are essential for binding. The subsequent experimental validation of the relative importance of residues in context provides a comprehensive understanding of the molecular recognition event between PfAMA1 and PfRON2, specifically in the post-binding stage. This insight could potentially open up new avenues to drug discovery against malaria.

Design, Synthesis, Biological Evaluation and Molecular Modeling Studies of Novel Naphthoquinone‐Triazole Hybrids as Potential FGFR1 Tyrosine Kinase Inhibitors

A new series of 1,4‐naphthoquinone‐triazole derivatives 5‐21 were synthesized using nucleophilic substitution and CuAAC reactions. The compounds were investigated for their cytotoxic activities against four cancer cell lines (i.e., HuCCA‐1, T47D, MOLT‐3, and HepG2) as well as a normal cell line (Vero). Most of the compounds showed active cytotoxic effects against all tested cancer cells without cytotoxicity to the normal cell. Particularly, compound 20 showed promising activity against the T47D with comparable effect to that of the known drug, doxorubicin. Compounds 16 and 21 exhibited the greatest FGFR1 inhibitory potency with nanomolar IC50 values of 1.31 ± 0.42 and 3.17 ± 0.27 nM, respectively. Interestingly, the derivative 16 showed comparable inhibitory potency with the known FGFR1 inhibitor, AZD4547. Molecular docking and molecular dynamics were conducted and revealed that both compounds could occupy within the same binding pocket of the target FGFR1 and shared common interacting key amino acids residues (i.e., Leu484, Val492, and Leu630) with those of inhibitor, AZD4547. The simulations also suggested that the naphthoquinone‐triazole skeleton was found to be a promising structural characteristic essential for effective inhibition of FGFR1. Additionally, the drug‐likeness prediction suggested that these compounds (16 and 21) are drug‐like molecules with possibility for further development.

Parkinson-like wild-type superoxide dismutase 1 pathology induces nigral dopamine neuron degeneration in a novel murine model

Atypical wild-type superoxide dismutase 1 (SOD1) protein misfolding and deposition occurs specifically within the degenerating substantia nigra pars compacta (SNc) in Parkinson disease. Mechanisms driving the formation of this pathology and relationship with SNc dopamine neuron health are yet to be fully understood. We applied proteomic mass spectrometry and synchrotron-based biometal quantification to post-mortem brain tissues from the SNc of Parkinson disease patients and age-matched controls to uncover key factors underlying the formation of wild-type SOD1 pathology in this disorder. We also engineered two of these factors - brain copper deficiency and upregulated SOD1 protein levels - into a novel mouse strain, termed the SOCK mouse, to verify their involvement in the development of Parkinson-like wild-type SOD1 pathology and their impact on dopamine neuron health. Soluble SOD1 protein in the degenerating Parkinson disease SNc exhibited altered post-translational modifications, which may underlie changes to the enzymatic activity and aggregation of the protein in this region. These include decreased copper binding, dysregulation of physiological glycosylation, and atypical oxidation and glycation of key SOD1 amino acid residues. We demonstrated that the biochemical profile introduced in SOCK mice promotes the same post-translational modifications and the development of Parkinson-like wild-type SOD1 pathology in the midbrain and cortex. This pathology accumulates progressively with age and is accompanied by nigrostriatal degeneration and dysfunction, which occur in the absence of α-synuclein deposition. These mice do not exhibit weight loss nor spinal cord motor neuron degeneration, distinguishing them from transgenic mutant SOD1 mouse models. This study provides the first in vivo evidence that mismetallation and altered post-translational modifications precipitates wild-type SOD1 misfolding, dysfunction, and deposition in the Parkinson disease brain, which may contribute to SNc dopamine neuron degeneration. Our data position this pathology as a novel drug target for this disorder, with a particular focus on therapies capable of correcting alterations to SOD1 post-translational modifications.

Salivary Metabolomics as a Diagnostic Tool: Distinct Metabolic Profiles Across Orofacial Pain Subtypes

Orofacial pain (OFP) includes chronic pain conditions categorized into musculoskeletal (MS), neurovascular (NV), and neuropathic (NP) pain types, encompassing temporomandibular disorders (TMD), migraines, trigeminal neuralgia (TN), post-traumatic neuropathies, and burning mouth syndrome (BMS). These conditions significantly affect quality of life; yet, their underlying metabolic disruptions remain inadequately explored. Salivary metabolomics provides a non-invasive method to investigate biochemical alterations associated with OFP subtypes. This study aimed to identify pain-specific salivary metabolites across chronic OFP types and examine their correlations with clinical characteristics. Saliva samples from 63 OFP patients (TMD, migraines, TN, post-traumatic neuropathies, BMS) and 37 pain-free controls were analyzed using liquid chromatography–mass spectrometry (LC-MS) targeting 28 metabolites linked to pain. Statistical analyses determined significant metabolite changes and associations with pain subtypes and patient characteristics. Among the 28 analyzed metabolites, 18 showed significant differences between OFP patients and controls. Key amino acids, including DL-glutamic acid, DL-aspartic acid, DL-citrulline, spermidine, and DL-ornithine, were significantly elevated in MS, NV, and NP pain types compared to controls. Additionally, DL-glutamine, DL-valine, and DL-phenylalanine were distinctively elevated in TMD and migraine patients. BMS displayed fewer alterations, with significantly lower levels of DL-proline, DL-tryptophan, DL-glutamic acid, DL-asparagine, and DL-aspartic acid compared to other pain types but elevated spermidine levels relative to controls. Salivary metabolomics revealed distinct metabolic alterations in OFP subtypes, providing insights into potential biomarkers for diagnosis and monitoring. These findings offer a foundation for personalized approaches in OFP management, although further research is required to validate and expand these results.

Structural and biological investigations of Fe(III) and Co(II) complexes with tryptophan and 2,2'-bipyridine: implications for antibacterial and antifungal applications

Abstract This study focused on the design and stractural characterization of two new transition metal complexes derived from Tryptophan (Trp) and 2,2'-bipyridine (Bip), coordinated with Iron(III) (FeTrpBip) and Cobalt(II) (CoTrpBip) ions. Structural elucidation of these complexes was achieved using a range of advanced analytical techniques. Thermal analysis revealed the stability and decomposition behaviors of the complexes. The data indicated that both FeTrpBip and CoTrpBip exhibit octahedral coordination geometries, with the structural formulas identified as [Fe(Trp)(Bip)(Cl)2] and [Co(Trp)(Bip)(Cl)(H2O)], respectively. To support the experimental data, Density Functional Theory (DFT) calculations had been performed. These calculations confirmed the proposed structures and provided a detailed analysis of quantum chemical parameters, including HOMO–LUMO energies, molecular orbitals, and electronic distributions, which are important for understanding the complexes' reactivity. Further, extensive in vitro biological evaluations assessed the antifungal and antibacterial evaluation of the synthesized metal complexes. The bioassays demonstrated that both FeTrpBip and CoTrpBip displayed significantly enhanced bioactivity compared to the free ligands, indicating a synergistic effect of metal coordination on the biological efficacy of the ligands. Molecular docking studies were subsequently conducted to explore the mechanisms of action of these complexes at the molecular level, specifically targeting the E. coli FabH–CoA (PDB ID: 1HNJ). The FabH receptor, essential for fatty acid biosynthesis, was chosen to evaluate the antimicrobial potential of the complexes. Docking simulations provided valuable insights into binding affinities, interaction energies, and key amino acid residues involved in the binding process. The results from these extensive studies highlight the significant therapeutic potential of FeTrpBip and CoTrpBip complexes, positioning them as promising reagents for further development in medical science. The observed synergistic effects due to metal coordination underscore the potential for these complexes to advance antimicrobial therapies and address challenges associated with resistant strains.

Semi-rational engineering of 7β-hydroxysteroid dehydrogenase enhances forward reaction activity towards ursodeoxycholic acid synthesis.

Semi-rational engineering of 7β-hydroxysteroid dehydrogenase enhances forward reaction activity towards ursodeoxycholic acid synthesis.

De novo synthesis of hyaluronic acid with tailored molecular weights using a new hyaluronidase SthHL.

De novo synthesis of hyaluronic acid with tailored molecular weights using a new hyaluronidase SthHL.

In silico identification of a phosphate marine steroid from Indonesian marine compounds as a potential inhibitor of phosphatidylinositol mannosyltransferase (PimA) in Mycobacterium tuberculosis.

In silico identification of a phosphate marine steroid from Indonesian marine compounds as a potential inhibitor of phosphatidylinositol mannosyltransferase (PimA) in Mycobacterium tuberculosis.

AmyA contributes to the glycogen synthesis in Sulfolobus acidocaldarius.

AmyA contributes to the glycogen synthesis in Sulfolobus acidocaldarius.

In silico discovery of a novel potential allosteric PI3Kα inhibitor incorporating 2-oxopropyl urea targeting head and neck squamous cell carcinoma

Head and neck squamous cell carcinoma (HNSCC) is the most common head and neck cancer and highly aggressive and heterogeneous. Targeted therapy is still the main treatment method used in clinic due to lower side effect and personalized medication. In order to discover novel and effective drugs with low side effect against HNSCC, we analyzed the genes related to HNSCC, and found that PIK3CA was highly expressed in tumor tissues and often experienced mutations, leading to excessive activation of phosphoinositide 3-kinase alpha (PI3Kα), promoting the development of HNSCC. The allosteric PI3Kα inhibitor STX-478 inhibits the growth of tumor with hotspot mutations in PI3Kα and shows prominent efficacy on the treatment of human HNSCC xenografts without displaying the metabolic dysfunction observed in Alpelisib. These mutations open the allosteric site more readily, increasing the selectivity of STX-478 for mutant PI3Kα. STX-478 cleverly avoids the side effect of ATP competitive PI3Kα inhibitors. So, the structure of STX-478 was optimized based on the interaction mechanism between STX-478 and PI3Kα. Then, virtual screening, binding mode research, target verification, physical and chemical properties, pharmacokinetic properties and stabilities of ligand-PI3Kα complexes were evaluated by computer technologies (scaffold hopping, cdocker, SuperPred, SwissTarget prediction, Lipinski’s rule of five, ADMET and MD simulation). Finally, J-53 (2-oxopropyl urea compound) with excellent properties was selected. J-53 not only formed H-bonds with key amino acids, but its unique -C(O)CH3 could also form H-bonds with ILE1019, making it more stably bound to PI3Kα and contributing to its activity. After the SciFinder verification, J-53 with novel structure had the value of further study. This study suggested that J-53 could be used as potential inhibitors of PI3Kα, and provides valuable information for the subsequent drug discovery of allosteric PI3Kα inhibitors.Graphical

A single residue in domain II of envelope protein of yellow fever virus is critical for neutralization sensitivity.

The YF-17D vaccine has been used to prevent YF disease. However, recent strains belonging to the SAI displayed reduced sensitivity to the antibodies produced by vaccination, raising concerns about potential future outbreaks. To identify potential amino acid residues responsible for the decreased neutralizing activity of YF-17D-vaccinated sera, we conducted a screening and generated recombinant viruses with amino acid changes specific to the SAI in EDII using the YF-17D genome as a genetic backbone. We found the A83E mutation played a key role in reducing neutralizing sensitivity to YF-17D-vaccinated mouse sera. Importantly, the A83E mutant maintained a comparable attenuation phenotype to YF-17D but elicited enhanced neutralization activity and conferred protection in mice. Together, we identify a key amino acid residue responsible for the neutralization escape of SAI YFV isolates. We propose that this substitution could act as a target for developing an updated YF-17D vaccine.

Metabolomic profiling of patients with gestational diabetes mellitus

Relevance. Pregnancy in women with carbohydrate metabolism disorders is classified as a high-risk group for early reproductive losses, obstetric complications, and perinatal morbidity, despite undeniable progress in understanding the pathogenesis, timely correction, and prevention of complications in this cohort of women. Aim: To study the metabolomic profile of amino acids in women with gestational diabetes mellitus (GDM) to identify potential biomarkers for predicting obstetric and perinatal complications. Materials and Methods: The study involved 50 pregnant women in the third trimester: 25 with GDM (GDM group) and 25 without GDM (control group). The concentration of 26 amino acids in a morning urine sample was determined for all participants, with analysis performed on the Agilent 1200 HPLC chromatographic system. Statistical processing of the data was carried out using IBM SPSS v. 26. Results: Three significant metabolites — valine, lysine, and glutamine — were identified. In GDM group, lysine levels were 5.3 times lower, valine 2 times lower, and glutamine 1.61 times lower compared to the control group. These changes may reflect GDM pathogenesis and predict complications. Conclusion: To enhance the effectiveness of diagnosis, individual management of pregnant women, and prevention of complications in women with GDM, it is important to consider the reduction in concentrations of key amino acids such as valine, lysine, and glutamine, which can serve as predictors of carbohydrate metabolism disorders.

Insights into the mechanism of substrate specificity in a novel PL15_3 subfamily oligo-alginate lyase VBAly15A.

Alginate is a major component of brown algae cell walls and can be degraded via β-elimination by alginate lyases. These enzymes are classified into polysaccharide lyases and oligo-alginate lyases (Oals), with Oals mainly represented by the PL15 and PL17 families. Unlike PL17 Oals, which are widely present in alginate-degrading microorganisms, PL15 enzymes are only identified in a limited number of microorganisms, and their biochemical characteristics remain poorly understood. In this research, a novel PL15 alginate lyase, VBAly15A, from the marine bacterium, Vibrio sp. B1Z05, was identified and characterized. It belongs to a new PL15_3 subfamily and exhibits high activity toward polyM substrates. VBAly15A is thermostable in medium temperatures, tolerant to alkaline up to 11.0, and polyM-specific Oal, and it can first degrade alginate polymers into disaccharides and subsequently catalyze disaccharides into monomers via an exolytic mode. Site-directed mutagenesis showed that Arg114, Tyr470, and Arg110 in the active groove are essential for the stable binding of the substrate. In addition, the amino acid His226 in VBAly15A, previously suggested to act as a catalytic base, is not essential for catalysis, whereas Tyr280, previously proposed to act as a catalytic acid, is required for enzyme activity. Structural bioinformatic and biochemical analyses revealed that His226 functions as a catalytic base, specifically abstracting protons from G-type substrates, while Tyr280 acts as both a catalytic acid and a base. This catalytic mechanism is likely conserved in PL15 family alginate lyases.IMPORTANCEAlginate, as a renewable resource for sustainability, has great application prospects. In addition to polysaccharide lyases, Oals are critical for the full degradation of alginate, a key prerequisite for biorefinery. So far, most identified and well-characterized Oals belong to the PL17 family. However, the catalytic mechanism of PL15 Oals is limited, and even the catalytic base and acid are not fully elucidated. The significance of this study lies in discovering and characterizing a novel Oal VBAly15A that divides into a new PL15 subfamily, PL15_3. Not only are key amino acid residues involved in enzyme activity identified, but residues acting as the catalytic base and acid are also demonstrated. The distance of the catalytic residues His and Tyr to the C5 proton of the sugar ring determines the substrate specificity. Therefore, this work provides new insights into the mechanism of substrate specificity in alginate lyases.

Molecular analysis of influenza A(H3N2) in a remote tropical island during 2014-2019 to identify the frequency of introduction and local circulation.

Molecular analysis of influenza A(H3N2) in a remote tropical island during 2014-2019 to identify the frequency of introduction and local circulation.

Synthesis, Physicochemical Properties and Anti-Fungal Activities of New Meso-Arylporphyrins

In this work, we describe the synthesis of three new meso-arylporphyrins, named meso-tetrakis [4-(nicotinoyloxy)phenyl] porphyrin (H2TNPP), meso-tetrakis [4-(picolinoyloxy)phenyl] porphyrin (H2TPPP), and meso-tetrakis [4-(isonicotinoyloxy) phenyl] porphyrin (H2TIPP). These new synthesized meso-arylporphyrins are characterized using spectroscopic analysis: Fourier Transform Infrared Spectroscopy (FTIR) and One-dimensional Nuclear Magnetic Resonance (1D NMR), and mass spectrometry (MS). The photophysical studies (UV–visible absorption, singlet oxygen (1O2) luminescence, and fluorescence emissions) demonstrate their potential uses as photosensitizers (PSs) in photodynamic therapy (PDT) applications. An in vitro investigation of the anti-fungal activity of H2TNPP, H2TPPP, and H2TIPP against Candida (C.) species (C. albicans, C. glabrata, and C. tropicalis) reveals that their minimum inhibitory concentration (MIC) values ranged from 1.25 to 5 mg/mL. In addition, their in vitro anti-fungal susceptibilities against three dermatophyte clinical isolates (Trichophyton rubrum, Microsporum canis, and Trichophyton mentagrophytes) are also evaluated and they demonstrate good anti-fungal activities. A molecular docking study of these meso-arylporphyrins as anti-fungal agents against C. tropicalis extracellular aspartic proteinases, Protein data Bank in Europe (PDBe code: 1J71) and Trichophyton rubrum Sialidases (PDBe code: 7P1D) underlines the possible interactions of H2TNPP, H2TPPP, and H2TIPP with the key amino acid residues of these fungal target proteins.

Mechanistic study of α-mangostin derivatives as potent α-glucosidase inhibitors.

α-Glucosidase inhibitors (AGIs) are pharmacological agents commonly used to manage postprandial hyperglycemia associated with type 2 diabetes mellitus (T2DM). Developing novel, potent AGIs remains a significant area of research. In this study, we investigated a series of derivatives of the natural product from α-mangostin as potential AGIs. A combined experimental and computational approach was employed to characterize promising compounds with potent α-glucosidase inhibitory activity. We found that α-mangostin (AM) and its derivatives (AM1 - 3) exhibited micromolar range α-glucosidase inhibition (IC50 ranging from 15.14 to 67.81µM), surpassing the known drug acarbose (IC50 of 197.09µM). Among the derivatives, AM1 exhibited the most promising α-glucosidase inhibition, displaying competitive inhibition kinetics with a Ki value of 47.04µM. Molecular docking and molecular dynamics (MD) simulations provided mechanistic insights into the binding interactions between AM1 and the α-glucosidase active site. AM1 was observed to form hydrogen bonds and hydrophobic interactions with key amino acid residues within the enzyme's active site. The introduction of amine groups in compound AM1 enhanced activity compared to AM, the parent compound. This study highlights the potential of α-mangostin derivatives as potent AGIs. The identified lead compound, AM1, warrants further investigation to assess its efficacy and safety in managing T2DM.

Discovery of a selective PI3Kα inhibitor via structure-based virtual screening for targeted colorectal cancer therapy

Colorectal cancer (CRC) remains a leading cause of cancer-related mortality globally, driving an urgent need for effective therapies. A promising avenue of research focuses on the PI3K/AKT/mTOR signalling pathway, which is frequently disrupted by mutations in the PI3Kα subunit. Our cutting-edge study employed a structure-based virtual screening of ∼3000 compounds, leading to the discovery of F0608-0019, a highly potent and selective PI3Kα inhibitor. F0608-0019 demonstrated remarkable efficacy in suppressing HCT116 colorectal cancer cell proliferation, with an IC50 of 12.14 µM, while maintaining high selectivity by minimising activity against other PI3K isoforms. Advanced molecular dynamics simulations highlighted the stability of F0608-0019’s binding interactions with key amino acids, such as TRP:780, ILE:932, and VAL:850, which are critical for its targeted action. These exciting findings reveal F0608-0019 as a leading candidate for innovative CRC therapies that selectively target PI3Kα dysregulation, offering promising new possibilities for effective CRC treatment.

Identification and Validation of Key Amino Acids in IgE Linear Epitopes of β-Lactoglobulin: Comparison of Recognition Patterns of Chinese Bovine Milk-Allergic Sera with Different Symptoms.

β-lactoglobulin (BLG) is the primary allergen in bovine milk allergy. The identification of key amino acids in the BLG epitopes has not been comprehensive due to differences in identification methods, patient symptoms, and population characteristics. In this study, bioinformatics predictions were conducted for key amino acids based on two potential IgE linear epitopes in BLG. Then, the peptide AA30-43 was confirmed as an IgE linear epitope through alanine scanning mutagenesis and peptide microarray assays, with four key amino acids (A34, A37, R40, and V41) common to different symptoms being identified. Moreover, symptom-specific key amino acids were identified. Serine (S30) and aspartic acid (D33) are the key amino acids for cutaneous allergy, while food allergy sera showed a preference for recognizing leucines in different positions (L31 and L39). Additionally, mutant peptides (R40, V41, L39, and D33) showed an obvious decrease in digestive stability compared with the epitope. Finally, the results of the KU812 cell degranulation model validated the critical role of the amino acids in allergenicity. These findings offer significant advantages for advancing both immune tolerance therapies and hypoallergenic milk product development, which hold significant implications for further research, prevention, and treatment of bovine milk allergy.

A Study of Variation in the Major Phenolic Acid Components of Dandelions Across Different Regions, and the Potential Molecular Mechanisms of Their Anti-Inflammatory Activity

This study explores the variation in the content of major phenolic acid components in dandelions from different regions, and the potential molecular mechanisms underlying their anti-inflammatory activity. High-performance liquid chromatography (HPLC) was employed to analyze dandelion leaves collected from four different regions in Hebei Province across eight harvest periods. The results indicated that chlorogenic acid had the highest content (0.334–1.963%), suggesting that this could be a key evaluation index for dandelion leaf harvesting. Further molecular docking and molecular dynamics simulations revealed that chlorogenic acid, caffeic acid, and chicoric acid could competitively bind to the key amino acid residues (e.g., PHE-151, ILE-117) of the MD-2 protein, thereby preventing the insertion of lipopolysaccharides (LPSs) and inhibiting the formation of the TLR4/MD-2 complex, which elucidates their potential anti-inflammatory mechanism. Moreover, environmental factors significantly influenced the accumulation of phenolic acids in dandelions, with temperature, precipitation, soil pH, and altitude showing correlations with the content variation of major phenolic acids. These findings provide a scientific basis for determining the optimal harvesting period of dandelion leaves, and offer new insights into the anti-inflammatory mechanisms of phenolic acids.

Comprehensive Analysis and Comparison of Amino Acid Levels in Cerebrospinal Fluid and Plasma of Children with Leukemia by the LC-MS Technique

This study aimed to develop and optimize an analytical method for profiling 21 amino acids in cerebrospinal fluid and plasma, addressing the need for improved diagnostic tools in leukemia research. Using high-performance liquid chromatography coupled with electrospray ionization mass spectrometry, the method achieved enhanced resolution, sensitivity, and specificity. Rigorous sample preparation, including liquid–liquid extraction, ensured high recovery rates, while validation confirmed the method’s accuracy and reproducibility. Clinical application in pediatric leukemia patients revealed significant variations in amino acid concentrations across treatment stages, providing insights into disease progression and therapeutic response. Statistical analysis with IBM SPSS Statistics 25 compared amino acid levels in patients to healthy controls, identifying distinct patterns on day 1, day 15, and day 33 of treatment. Correlation analysis highlighted relationships between amino acid levels and factors such as treatment duration, sex, age, and blood test results. Key amino acids, including proline, leucine, and hydroxyproline, emerged as significant predictors of white blood cell count, effectively distinguishing between patient and control groups. This method demonstrates robust potential for broader leukemia research applications, pending further validation on larger cohorts.