RESUMEN

Background: Atopic dermatitis (AD) is a chronic inflammatory skin condition that has a significant impact on quality of life. The immune response and allergy symptoms in AD are triggered by the recognition of specific allergens by IgE antibodies. Cross-reactivity can lead to auto-IgE responses, potentially worsening AD symptoms. Our research aimed to enhance our understanding of allergenic sources, including A. fumigatus, and their role in AD. We focused on molecular mimicry between human AQP3 and A. fumigatus aquaporin. Methods: In our in-silico analysis, we compared the amino acid sequences of human aquaporin 3 (AQP3) and A. fumigatus aquaporin with 25 aquaporins from various allergenic sources, sourced from the UniProt and NCBI databases. Phylogenetic relationship analysis and homology-based modeling were conducted. We identified conserved antigenic regions located within the 3D structures. Results: The global identity levels among the studied aquaporins averaged 32.6%. One antigenic site exhibited a remarkable local region, with a conserved identity of 71.4%. We categorized the aquaporins into five monophyletic clades (A-E), with group B showing the highest identity (95%), including six mammalian aquaporins, including AQP3. When comparing A. fumigatus aquaporins, the highest identity was observed with Malassezia sympodialis at 35%. Both human and A. fumigatus aquaporins have three linear and three discontinuous epitopes. Conclusions: We identified potential linear and conformational epitopes of AQP3, indicating a possible molecular mimicry between humans and A. fumigatus aquaporins. This suggests autoreactivity and potential cross-reactivity, although further validation using in vitro and in vivo experiments is required.

Asunto(s)

Acuaporina 3 , Acuaporinas , Aspergillus fumigatus , Simulación por Computador , Imitación Molecular , Filogenia , Humanos , Aspergillus fumigatus/inmunología , Aspergillus fumigatus/metabolismo , Acuaporina 3/metabolismo , Acuaporinas/metabolismo , Acuaporinas/química , Acuaporinas/genética , Secuencia de Aminoácidos , Alérgenos/inmunología , Alérgenos/metabolismo , Hipersensibilidad/inmunología , Hipersensibilidad/microbiología , Modelos Moleculares , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/inmunología

RESUMEN

Peptides are receiving significant attention in pharmaceutical sciences due to their applications as anti-inflammatory drugs; however, many aspects of their interactions and mechanisms at the molecular level are not well-known. This work explores the molecular structure of two peptides-(i) cysteine (Cys)-asparagine (Asn)-serine (Ser) (CNS) as a molecule in the gas phase and solvated in water in zwitterion form, and (ii) the crystal structure of the dipeptide serine-asparagine (SN), a reliable peptide indication whose experimental cell parameters are well known. A search was performed by means of atomistic calculations based on density functional theory (DFT). These calculations matched the experimental crystal structure of SN, validating the CNS results and useful for assignments of our experimental spectroscopic IR bands. Our calculations also explore the intercalation of CNS into the interlayer space of montmorillonite (MNT). Our quantum mechanical calculations show that the conformations of these peptides change significantly during intercalation into the confined interlayer space of MNT. This intercalation is energetically favorable, indicating that this process can be a useful preparation for therapeutic anti-inflammatory applications and showing high stability and controlled release processes.

Asunto(s)

Antiinflamatorios , Bentonita , Cisteína , Teoría Funcional de la Densidad , Serina , Bentonita/química , Antiinflamatorios/química , Antiinflamatorios/farmacología , Cisteína/química , Serina/química , Asparagina/química , Modelos Moleculares , Péptidos/química , Sustancias Intercalantes/química

RESUMEN

Deep learning methods, trained on the increasing set of available protein 3D structures and sequences, have substantially impacted the protein modeling and design field. These advancements have facilitated the creation of novel proteins, or the optimization of existing ones designed for specific functions, such as binding a target protein. Despite the demonstrated potential of such approaches in designing general protein binders, their application in designing immunotherapeutics remains relatively underexplored. A relevant application is the design of T cell receptors (TCRs). Given the crucial role of T cells in mediating immune responses, redirecting these cells to tumor or infected target cells through the engineering of TCRs has shown promising results in treating diseases, especially cancer. However, the computational design of TCR interactions presents challenges for current physics-based methods, particularly due to the unique natural characteristics of these interfaces, such as low affinity and cross-reactivity. For this reason, in this study, we explored the potential of two structure-based deep learning protein design methods, ProteinMPNN and ESM-IF1, in designing fixed-backbone TCRs for binding target antigenic peptides presented by the MHC through different design scenarios. To evaluate TCR designs, we employed a comprehensive set of sequence- and structure-based metrics, highlighting the benefits of these methods in comparison to classical physics-based design methods and identifying deficiencies for improvement.

Asunto(s)

Biología Computacional , Aprendizaje Profundo , Receptores de Antígenos de Linfocitos T , Receptores de Antígenos de Linfocitos T/inmunología , Receptores de Antígenos de Linfocitos T/química , Receptores de Antígenos de Linfocitos T/metabolismo , Biología Computacional/métodos , Humanos , Ingeniería de Proteínas/métodos , Modelos Moleculares , Conformación Proteica , Unión Proteica

RESUMEN

CONTEXT: Alzheimer's disease (AD) is the leading cause of dementia around the world, totaling about 55 million cases, with an estimated growth to 74.7 million cases in 2030, which makes its treatment widely desired. Several studies and strategies are being developed considering the main theories regarding its origin since it is not yet fully understood. Among these strategies, the 5-HT6 receptor antagonism emerges as an auspicious and viable symptomatic treatment approach for AD. The 5-HT6 receptor belongs to the G protein-coupled receptor (GPCR) family and is closely implicated in memory loss processes. As a serotonin receptor, it plays an important role in cognitive function. Consequently, targeting this receptor presents a compelling therapeutic opportunity. By employing antagonists to block its activity, the 5-HT6 receptor's functions can be effectively modulated, leading to potential improvements in cognition and memory. METHODS: Addressing this challenge, our research explored a promising avenue in drug discovery for AD, employing Artificial Neural Networks-Quantitative Structure-Activity Relationship (ANN-QSAR) models. These models have demonstrated great potential in predicting the biological activity of compounds based on their molecular structures. By harnessing the capabilities of machine learning and computational chemistry, we aimed to create a systematic approach for analyzing and forecasting the activity of potential drug candidates, thus streamlining the drug discovery process. We assembled a diverse set of compounds targeting this receptor and utilized density functional theory (DFT) calculations to extract essential molecular descriptors, effectively representing the structural features of the compounds. Subsequently, these molecular descriptors served as input for training the ANN-QSAR models alongside corresponding biological activity data, enabling us to predict the potential efficacy of novel compounds as 5-hydroxytryptamine receptor 6 (5-HT6) antagonists. Through extensive analysis and validation of ANN-QSAR models, we identified eight new promising compounds with therapeutic potential against AD.

Asunto(s)

Enfermedad de Alzheimer , Diseño de Fármacos , Relación Estructura-Actividad Cuantitativa , Receptores de Serotonina , Antagonistas de la Serotonina , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/metabolismo , Receptores de Serotonina/metabolismo , Receptores de Serotonina/química , Humanos , Antagonistas de la Serotonina/química , Antagonistas de la Serotonina/farmacología , Antagonistas de la Serotonina/uso terapéutico , Redes Neurales de la Computación , Modelos Moleculares

RESUMEN

Selective recognition and sensing of catecholamine-based neurotransmitters by fluorescent synthetic receptors capable of operating in pure water is a central topic of modern supramolecular chemistry that impacts biological and analytical chemistry. Despite advances achieved in the recognition of some neurotransmitters such as dopamine, little effort has been invested in the optical recognition of other neurotransmitters of paramount importance in biochemistry and medicinal chemistry such as the drug L-dihydroxy-phenylalanine (levodopa). Herein, a cationic Cu(II)-terpyridine complex bearing an intramolecular fluorescent quinolinium ring covalently linked to phenylboronic acid (CuL1) was synthesized, structurally described by single-crystal X-ray diffraction and studied in-depth as a fluorescent receptor for neurotransmitters in water. The complex CuL1 was designed to act as a receptor for levodopa through two Lewis acids of different natures (Cu(II) and B atoms) as cooperative binding points. The receptor CuL1 was found to have a strongly acidified -B(OH)2 group (pKa = 6.2) and exceptionally high affinity for levodopa (K = 4.8 × 106 M-1) with selectivity over other related neurotransmitters such as dopamine, epinephrine, norepinephrine and nucleosides in the micromolar concentration range at physiological pH. Such levodopa affinity/selectivity for a boronic acid-based receptor in water is still rare. On the basis of spectroscopic tools (11B NMR, UV-vis, EPR, and fluorescence), high-resolution ESI-MS, crystal structure, and DFT calculations, the interaction mode of CuL1 with levodopa is proposed in a 1 : 1 model using two-point recognition involving a boronate-catechol esterification and a coordination bond Cu(II)-carboxylate. Furthermore, a visual sensing ensemble was constructed using CuL1 and the commercial fluorescent dye eosin Y. Levodopa is efficiently detected by the displacement of the eosin Y bound to the Cu(II)-receptor, monitoring its green emission. The use of Cu(II)-boronate complexes for fast and selective neurotransmitter sensing was unexplored until now.

Asunto(s)

Ácidos Borónicos , Complejos de Coordinación , Cobre , Agua , Ácidos Borónicos/química , Agua/química , Cobre/química , Complejos de Coordinación/química , Complejos de Coordinación/síntesis química , Modelos Moleculares , Levodopa/química , Estructura Molecular , Sitios de Unión

RESUMEN

Four afatinib derivatives were designed and modeled. These derivatives were compared to the known tyrosine-kinase inhibitors in treating Chronic Myeloid Leukemia, i.e., imatinib and ponatinib. The molecules were evaluated through computational methods, including docking studies, the non-covalent interaction index, Electron Localization and Fukui Functions, in silico ADMET analysis, QTAIM, and Heat Map analysis. The AFA(IV) candidate significantly increases the score value compared to afatinib. Furthermore, AFA(IV) was shown to be relatively similar to the ponatinib profile when evaluating a range of molecular descriptors. The addition of a methylpiperazine ring seems to be well distributed in the structure of afatinib when targeting the BCR-ABL enzyme, providing an important hydrogen bond interaction with the Asp381 residue of the DFG-switch of BCR-ABL active site residue and the AFA(IV) new chemical entities. Finally, in silico toxicity predictions show a favorable index, with some molecules presenting the loss of the irritant properties associated with afatinib in theoretical predictions.

Asunto(s)

Afatinib , Proteínas de Fusión bcr-abl , Simulación del Acoplamiento Molecular , Inhibidores de Proteínas Quinasas , Proteínas de Fusión bcr-abl/antagonistas & inhibidores , Proteínas de Fusión bcr-abl/genética , Proteínas de Fusión bcr-abl/química , Afatinib/química , Afatinib/farmacología , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Humanos , Modelos Moleculares , Simulación por Computador , Mutación , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Enlace de Hidrógeno , Antineoplásicos/química , Antineoplásicos/farmacología , Imidazoles/química , Imidazoles/farmacología , Piridazinas

RESUMEN

Fold-switching enables metamorphic proteins to reversibly interconvert between two highly dissimilar native states to regulate their protein functions. While about 100 proteins have been identified to undergo fold-switching, unveiling the key residues behind this mechanism for each protein remains challenging. Reasoning that fold-switching in proteins is driven by dynamic changes in local energetic frustration, we combined fold-switching simulations generated using simplified structure-based models with frustration analysis to identify key residues involved in this process based on the change in the density of minimally frustrated contacts during refolding. Using this approach to analyze the fold-switch of the bacterial transcription factor RfaH, we identified 20 residues that significantly change their frustration during its fold-switch, some of which have been experimentally and computationally reported in previous works. Our approach, which we developed as an additional module for the FrustratometeR package, highlights the role of local frustration dynamics in protein fold-switching and offers a robust tool to enhance our understanding of other proteins with significant conformational shifts.

Asunto(s)

Proteínas de Escherichia coli , Pliegue de Proteína , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Transactivadores/química , Transactivadores/metabolismo , Transactivadores/genética , Simulación de Dinámica Molecular , Factores de Elongación de Péptidos/química , Factores de Elongación de Péptidos/metabolismo , Modelos Moleculares , Conformación Proteica , Termodinámica

RESUMEN

Aminoglycosides are essential antibiotics used to treat severe infections caused mainly by Gram-negative bacteria. Gentamicin is an aminoglycoside and, despite its toxicity, is clinically used to treat several pulmonary and urinary infections. The commercial form of gentamicin is a mixture of five compounds with minor differences in the methylation of one of their aminosugars. In the case of two compounds, gentamicin C2 and C2a, the only difference is the stereochemistry of the methyl group attached to C-6'. GenB2 is the enzyme responsible for this epimerization and is one of the four PLP-dependent enzymes encoded by the gentamicin biosynthetic gene cluster. Herein, we have determined the structure of GenB2 in its holo form in complex with PMP and also in the ternary complex with gentamicin X2 and G418, two substrate analogues. Based on the structural analysis, we were able to identify the structural basis for the catalytic mechanism of this enzyme, which was also studied by site-directed mutagenesis. Unprecedently, GenB2 is a PLP-dependent enzyme from fold I, which is able to catalyze an epimerization but with a mechanism distinct from that of fold III PLP-dependent epimerases using a cysteine residue near the N-terminus. The substitution of this cysteine residue for serine or alanine completely abolished the epimerase function of the enzyme, confirming its involvement. This study not only contributes to the understanding of the enzymology of gentamicin biosynthesis but also provides valuable details for exploring the enzymatic production of new aminoglycoside derivatives.

Asunto(s)

Gentamicinas , Gentamicinas/metabolismo , Gentamicinas/biosíntesis , Gentamicinas/química , Antibacterianos/química , Antibacterianos/biosíntesis , Antibacterianos/metabolismo , Racemasas y Epimerasas/metabolismo , Racemasas y Epimerasas/genética , Racemasas y Epimerasas/química , Modelos Moleculares , Cristalografía por Rayos X , Mutagénesis Sitio-Dirigida , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/química , Proteínas Bacterianas/genética

RESUMEN

Nerve agents are organophosphates (OPs) that act as potent inhibitors of acetylcholinesterase (AChE), the enzyme responsible for the hydrolysis of acetylcholine. After inhibition, a dealkylation reaction of the phosphorylated serine, known as the aging of AChE, can occur. When aged, reactivators of OP-inhibited AChE are no longer effective. Therefore, the realkylation of aged AChE may offer a pathway to reverse AChE aging. In this study, molecular modeling was conducted to propose new ligands as realkylators of aged AChE. We applied a methodology involving docking and quantum mechanics/molecular mechanics (QM/MM) calculations to evaluate the resurrection kinetic constants and ligand interactions with OP-aged AChE, comparing them to data found in the literature. The results obtained confirm that this method is suitable for predicting kinetic and thermodynamic parameters of ligands, which can be useful in the design and selection of new and more effective ligands for AChE realkylation.

Asunto(s)

Acetilcolinesterasa , Inhibidores de la Colinesterasa , Indolquinonas , Acetilcolinesterasa/química , Acetilcolinesterasa/metabolismo , Inhibidores de la Colinesterasa/química , Inhibidores de la Colinesterasa/farmacología , Cinética , Indolquinonas/química , Simulación del Acoplamiento Molecular , Ligandos , Termodinámica , Modelos Moleculares , Humanos , Simulación de Dinámica Molecular

RESUMEN

To ascertain the bioinorganic chemistry of metals conjugated with quinones, the complexes [Ag(ATV)(PPh3)2] (1), [Au(ATV)(PPh3)]·2H2O (2), and [Cu(ATV)(PPh3)2] (3) were synthesized by the coordination of the antimalarial naphthoquinone atovaquone (ATV) to the starting materials [Ag(PPh3)2]NO3, [Au(PPh3)Cl], and [Cu(PPh3)2NO3], respectively. These complexes were characterized by analytical and spectroscopical techniques. X-ray diffraction of single crystals precisely confirmed the coordination mode of ATV to the metals, which was monodentate or bidentate, depending on the metal center. Both coordination modes showed high stability in the solid state and in solution. All three complexes showed negative log D values at pH 5, but at pH 7.4, while complex 2 continued to have a negative log D value, complexes 1 and 3 displayed positive values, indicating a more hydrophilic character. ATV and complexes 1-3 could bind to ferriprotoporphyrin IX (FePPIX); however, only complexes 1-3 could inhibit ß-hematin crystal formation. Phenotype-based activity revealed that all three metal complexes are able to inhibit the growth of P. falciparum with potency and selectivity comparable to those of ATV, while the starting materials lack this activity. The outcomes of this chemical design may provide significant insights into structure-activity relationships for the development of new antimalarial agents.

Asunto(s)

Antimaláricos , Atovacuona , Complejos de Coordinación , Hemo , Plasmodium falciparum , Antimaláricos/farmacología , Antimaláricos/química , Antimaláricos/síntesis química , Plasmodium falciparum/efectos de los fármacos , Complejos de Coordinación/farmacología , Complejos de Coordinación/química , Complejos de Coordinación/síntesis química , Hemo/química , Atovacuona/farmacología , Atovacuona/química , Atovacuona/síntesis química , Estructura Molecular , Cobre/química , Cobre/farmacología , Plata/química , Plata/farmacología , Oro/química , Oro/farmacología , Fosfinas/química , Fosfinas/farmacología , Pruebas de Sensibilidad Parasitaria , Relación Estructura-Actividad , Modelos Moleculares , Humanos

RESUMEN

More and more attention has been paid to food safety. Due to the overuse and misuse of antibiotics, the problem of antibiotic residues in animal food is one of the important challenges to ensure food safety. The development of a feasible strategy to detect antibiotic residues in animal food has become desirable. In this paper, we creatively synthesize a water-stable fluorescence sensing material, namely, Co(Ⅱ)-Coordination polymer [Co2(CA) (L)0.5 (H2O)3] n (L = 1,4-bis(imidazole-1-ylmethyl) benzene, CA= Citric acid). The single crystal X-ray diffraction shows that it crystallizes in tetragonal space group I-4. It is worth mentioning that there exists the rare Co4(µ3-O)4 cubane cluster structure and Co8 cluster units. Those adjacent Co8 cluster units are connected into an infinite two-dimensional net structure by four flexible bridged L ligands. Finally, the Co(Ⅱ)-Coordination polymer (CP) further develops into the three-dimensional supramolecular structure via the hydrogen bonds of O-H⋯O and C-H⋯O. It could selectively detect the antibiotic-nitrofurantoin (NFT) residue by way of fluorescence quenching, Co-CP for the detection of NFT shows broad linearity from 0 to 200 µM, with a detection limit of 0.13 µM and strong anti-interference ability. It is used to detect the NFT residual of tap water and milk with a spiked recovery of 86.35-112.47 %.

Asunto(s)

Antibacterianos , Cobalto , Complejos de Coordinación , Colorantes Fluorescentes , Nitrofurantoína , Polímeros , Cobalto/química , Cobalto/análisis , Antibacterianos/análisis , Antibacterianos/química , Polímeros/química , Nitrofurantoína/análisis , Nitrofurantoína/química , Colorantes Fluorescentes/química , Complejos de Coordinación/química , Espectrometría de Fluorescencia/métodos , Animales , Leche/química , Modelos Moleculares , Contaminación de Alimentos/análisis , Fluorescencia

RESUMEN

Density Functional Theory (DFT) is extensively used in theoretical and computational chemistry to study molecular and crystal properties across diverse fields, including quantum chemistry, materials physics, catalysis, biochemistry, and surface science. Despite advances in DFT hardware and software for optimized geometries, achieving consensus in molecular structure comparisons with experimental counterparts remains a challenge. This difficulty is exacerbated by the lack of automated bond length comparison tools, resulting in labor-intensive and error-prone manual processes. To address these challenges, we propose MolGC, a Molecular Geometry Comparator algorithm that automates the comparison of optimized geometries from different theoretical levels. MolGC calculates the mean absolute error (MAE) of bond lengths by integrating data from various DFT software. It provides interactive and customizable visualization of geometries, enabling users to explore different views for enhanced analysis. In addition, it saves MAE computations for further analysis and offers a comprehensive statistical summary of the results. MolGC effectively addresses complex graph labeling challenges, ensuring accurate identification and categorization of bonds in diverse chemical structures. It achieves a 98.91% average rate in correct bond label assignments on an antibiotics dataset, showcasing its effectiveness for comparing molecular bond lengths across geometries of varying complexity and size. The executable file and software resources for running MolGC can be downloaded from https://github.com/AbimaelGP/MolGC/tree/main .

Asunto(s)

Algoritmos , Programas Informáticos , Teoría Funcional de la Densidad , Modelos Moleculares , Estructura Molecular

RESUMEN

Inulosucrases are enzymes capable of synthesizing inulin polymers using sucrose as the main substrate. The enzymatic activity relies on the catalytic triad within the active site and residues responsible for substrate recognition and orientation, termed carbohydrate-binding subsites. This study investigates the role of specific residues within the catalytic cavity of a truncated version of IslA4 in enzymatic catalysis. Mutants at residues S425, L499, A602, R618, F619, Y676, Y692, and R696 were constructed and characterized. Characterization results, and in silico structural comparison with other fructansucrases, reveal these residues' functional significance in catalysis. Residue S425 belongs to subsite -1; residues R618 and Y692 are part of subsite +1, and residue R696 belongs to subsites +1 and +2. Residues L499 and A602 are support residues; the former favors the formation of the fructosyl-enzyme intermediate, while the latter stabilizes the acid/base catalyst during catalysis. Residues Y676 and F619 may participate in stabilizing residues at -1/+1 subsites. This study represents the first comprehensive exploration of the structural determinants essential for enzymatic function in the inulosucrase of Leuconostoc citreum, and proposes the identity of residues involved in the -1 to +2 subsites.

Asunto(s)

Dominio Catalítico , Hexosiltransferasas , Leuconostoc , Leuconostoc/enzimología , Leuconostoc/genética , Hexosiltransferasas/genética , Hexosiltransferasas/metabolismo , Hexosiltransferasas/química , Modelos Moleculares , Especificidad por Sustrato , Secuencia de Aminoácidos , Cinética , Catálisis , Mutación , Proteínas Bacterianas

RESUMEN

To get a deeper understanding of the structural bases of the allosteric transition between T and R states of plant and bacterial phosphoenolpyruvate carboxylases (PEPCs), we obtained the first T-state crystal structures of the maize photosynthetic PEPC (ZmPEPC-C4) and exhaustively compared them with the previously reported R-state ZmPEPC-C4 and other T-state structures. We identified previously unrecognized significant conformational changes in the T state: that of the α8-α9 loop, which connects the two kinds of activator allosteric sites with the active site, the conversion of the α30 helix into a 310 helix, leading to the disorganization of the active site lid and activators allosteric sites, and the closure of the inhibitor allosteric-site lid. Additionally, we identified previously overlooked, highly conserved residues of potential interest in the allosteric transition, including two histidines whose protonation might stabilize the T state. The crystal structures reported here also suggest similar tetrameric quaternary arrangements of PEPC enzymes in the R and T states, and the location of the bicarbonate binding site, as well as the conformational changes required for the carboxylation step. Our findings and working hypothesis advance the understanding of the structural features of the allosteric PEPC enzymes and provide a foundation for future experiments.

Asunto(s)

Modelos Moleculares , Fosfoenolpiruvato Carboxilasa , Zea mays , Zea mays/enzimología , Zea mays/química , Regulación Alostérica , Cristalografía por Rayos X , Fosfoenolpiruvato Carboxilasa/química , Fosfoenolpiruvato Carboxilasa/metabolismo , Sitio Alostérico , Dominio Catalítico , Conformación Proteica , Secuencia de Aminoácidos

RESUMEN

Feline calicivirus (FCV), an important model for studying the biology of the Caliciviridae family, encodes the leader of the capsid (LC) protein, a viral factor known to induce apoptosis when expressed in a virus-free system. Our research has shown that the FCV LC protein forms disulfide bond-dependent homo-oligomers and exhibits intrinsic toxicity; however, it lacked a polybasic region and a transmembrane domain (TMD); thus, it was initially classified as a non-classical viroporin. The unique nature of the FCV LC protein, with no similarity to other proteins beyond the Vesivirus genus, has posed challenges for bioinformatic analysis reliant on sequence similarity. In this study, we continued characterizing the LC protein using the AlphaFold 2 and the recently released AlphaFold 3 artificial intelligence tools to predict the LC protein tertiary structure. We compared it to other molecular modeling algorithms, such as I-Tasser's QUARK, offering new insights into its putative TMD. Through exogenous interaction, we found that the recombinant LC protein associates with the CrFK plasmatic membrane and can permeate cell membranes in a disulfide bond-independent manner, suggesting that this interaction might occur through a TMD. Additionally, we examined its potential to activate the intrinsic apoptosis pathway in murine and human ovarian cancer cell lines, overexpressing survivin, an anti-apoptotic protein. All these results enhance our understanding of the LC protein's mechanism of action and suggest its role as a class-I viroporin.

Asunto(s)

Calicivirus Felino , Proteínas de la Cápside , Membrana Celular , Calicivirus Felino/metabolismo , Calicivirus Felino/genética , Proteínas de la Cápside/metabolismo , Proteínas de la Cápside/genética , Proteínas de la Cápside/química , Gatos , Animales , Membrana Celular/metabolismo , Modelos Moleculares , Línea Celular , Dominios Proteicos , Humanos , Apoptosis , Unión Proteica

RESUMEN

Drug repositioning is an important therapeutic strategy for treating breast cancer. Hsp90ß chaperone is an attractive target for inhibiting cell progression. Its structure has a disordered and flexible linker region between the N-terminal and central domains. Geldanamycin was the first Hsp90ß inhibitor to interact specifically at the N-terminal site. Owing to the toxicity of geldanamycin, we investigated the repositioning of ritonavir as an Hsp90ß inhibitor, taking advantage of its proven efficacy against cancer. In this study, we used molecular modeling techniques to analyze the contribution of the Hsp90ß linker region to the flexibility and interaction between the ligands geldanamycin, ritonavir, and Hsp90ß. Our findings indicate that the linker region is responsible for the fluctuation and overall protein motion without disturbing the interaction between the inhibitors and the N-terminus. We also found that ritonavir established similar interactions with the substrate ATP triphosphate, filling the same pharmacophore zone.

Asunto(s)

Benzoquinonas , Proteínas HSP90 de Choque Térmico , Lactamas Macrocíclicas , Ritonavir , Lactamas Macrocíclicas/farmacología , Lactamas Macrocíclicas/química , Ritonavir/química , Ritonavir/farmacología , Benzoquinonas/química , Benzoquinonas/farmacología , Benzoquinonas/metabolismo , Proteínas HSP90 de Choque Térmico/química , Proteínas HSP90 de Choque Térmico/metabolismo , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Humanos , Unión Proteica , Simulación de Dinámica Molecular , Simulación del Acoplamiento Molecular , Modelos Moleculares , Sitios de Unión , Adenosina Trifosfato/metabolismo , Adenosina Trifosfato/química

RESUMEN

The crystallographic structure of the FolB enzyme from Mycobacterium tuberculosis (MtFolB), complexed with its inhibitor 8-mercaptoguanine (8-MG), was elucidated at a resolution of 1.95 Å. A novel series of S8-functionalized 8-MG derivatives were synthesised and evaluated as in vitro inhibitors of dihydroneopterin aldolase (DHNA, EC 4.1.2.25) activity of MtFolB. These compounds exhibited IC50 values in the submicromolar range. Evaluation of the activity for five compounds indicated their inhibition mode and inhibition constants. Molecular docking analyses were performed to determine the enzyme-inhibitor intermolecular interactions and ligand conformations upon complex formation. The inhibitory activities of all compounds against the M. tuberculosis H37Rv strain were evaluated. Compound 3e exhibited a minimum inhibitory concentration in the micromolar range. Finally, Compound 3e showed no apparent toxicity in both HepG2 and Vero cells. The findings presented herein will advance the quest for novel, specific inhibitors targeting MtFolB, an attractive molecular target for TB drug development.

Asunto(s)

Aldehído-Liasas , Antituberculosos , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos , Pruebas de Sensibilidad Microbiana , Mycobacterium tuberculosis , Mycobacterium tuberculosis/efectos de los fármacos , Mycobacterium tuberculosis/enzimología , Antituberculosos/farmacología , Antituberculosos/síntesis química , Antituberculosos/química , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Humanos , Relación Estructura-Actividad , Aldehído-Liasas/antagonistas & inhibidores , Aldehído-Liasas/metabolismo , Aldehído-Liasas/química , Células Vero , Estructura Molecular , Cristalografía por Rayos X , Chlorocebus aethiops , Animales , Guanina/farmacología , Guanina/química , Guanina/análogos & derivados , Guanina/síntesis química , Simulación del Acoplamiento Molecular , Células Hep G2 , Modelos Moleculares

RESUMEN

Function and structure are strongly coupled in obligated oligomers such as Triosephosphate isomerase (TIM). In animals and fungi, TIM monomers are inactive and unstable. Previously, we used ancestral sequence reconstruction to study TIM evolution and found that before these lineages diverged, the last opisthokonta common ancestor of TIM (LOCATIM) was an obligated oligomer that resembles those of extant TIMs. Notably, calorimetric evidence indicated that ancestral TIM monomers are more structured than extant ones. To further increase confidence about the function, structure, and stability of the LOCATIM, in this work, we applied two different inference methodologies and the worst plausible case scenario for both of them, to infer four sequences of this ancestor and test the robustness of their physicochemical properties. The extensive biophysical characterization of the four reconstructed sequences of LOCATIM showed very similar hydrodynamic and spectroscopic properties, as well as ligand-binding energetics and catalytic parameters. Their 3D structures were also conserved. Although differences were observed in melting temperature, all LOCATIMs showed reversible urea-induced unfolding transitions, and for those that reached equilibrium, high conformational stability was estimated (ΔGTot = 40.6-46.2 kcal/mol). The stability of the inactive monomeric intermediates was also high (ΔGunf = 12.6-18.4 kcal/mol), resembling some protozoan TIMs rather than the unstable monomer observed in extant opisthokonts. A comparative analysis of the 3D structure of ancestral and extant TIMs shows a correlation between the higher stability of the ancestral monomers with the presence of several hydrogen bonds located in the "bottom" part of the barrel.

Asunto(s)

Triosa-Fosfato Isomerasa , Triosa-Fosfato Isomerasa/química , Triosa-Fosfato Isomerasa/genética , Triosa-Fosfato Isomerasa/metabolismo , Animales , Evolución Molecular , Multimerización de Proteína , Modelos Moleculares , Estabilidad de Enzimas

RESUMEN

Antioxidants agents play an essential role in the food industry for improving the oxidative stability of food products. In the last years, the search for new natural antioxidants has increased due to the potential high toxicity of chemical additives. Therefore, the synthesis and evaluation of the antioxidant activity in peptides is a field of current research. In this study, we performed a Quantitative Structure Activity Relationship analysis (QSAR) of cysteine-containing 19 dipeptides and 19 tripeptides. The main objective is to bring information on the relationship between the structure of peptides and their antioxidant activity. For this purpose, 1D and 2D molecular descriptors were calculated using the PaDEL software, which provides information about the structure, shape, size, charge, polarity, solubility and other aspects of the compounds. Different QSAR model for di- and tripeptides were developed. The statistic parameters for di-peptides model (R2train = 0.947 and R2test = 0.804) and for tripeptide models (R2train = 0.923 and R2test = 0.847) indicate that the generated models have high predictive capacity. Then, the influence of the cysteine position was analyzed predicting the antioxidant activity for new di- and tripeptides, and comparing them with glutathione. In dipeptides, excepting SC, TC and VC, the activity increases when cysteine is at the N-terminal position. For tripeptides, we observed a notable increase in activity when cysteine is placed in the N-terminal position.

Asunto(s)

Antioxidantes , Cisteína , Dipéptidos , Oligopéptidos , Relación Estructura-Actividad Cuantitativa , Cisteína/química , Antioxidantes/química , Antioxidantes/farmacología , Dipéptidos/química , Dipéptidos/farmacología , Oligopéptidos/química , Oligopéptidos/farmacología , Modelos Moleculares , Programas Informáticos

RESUMEN

Dienelactone hydrolase (DLH) is one of numerous hydrolytic enzymes with an α/ß-hydrolase fold, which catalyze the hydrolysis of dienelactone to maleylacetate. The DLHs share remarkably similar tertiary structures and a conserved arrangement of catalytic residues. This study presents the crystal structure and comprehensive functional characterization of a novel thermostable DLH from the bacterium Hydrogenobacter thermophilus (HtDLH). The crystal structure of the HtDLH, solved at a resolution of about 1.67 Å, exhibits a canonical α/ß-hydrolase fold formed by eight ß-sheet strands in the core, with one buried α-helix and six others exposed to the solvent. The structure also confirmed the conserved catalytic triad of DHLs formed by Cys121, Asp170, and His202 residues. The HtDLH forms stable homodimers in solution. Functional studies showed that HtDLH has the expected esterase activity over esters with short carbon chains, such as p-nitrophenyl acetate, reaching optimal activity at pH 7.5 and 70 °C. Furthermore, HtDLH maintains more than 50 % of its activity even after incubation at 90 °C for 16 h. Interestingly, HtDLH exhibits catalytic activity towards polyethylene terephthalate (PET) monomers, including bis-1,2-hydroxyethyl terephthalate (BHET) and 1-(2-hydroxyethyl) 4-methyl terephthalate, as well as other aliphatic and aromatic esters. These findings associated with the lack of activity on amorphous PET indicate that HtDLH has characteristic of a BHET-degrading enzyme. This work expands our understanding of enzyme families involved in PET degradation, providing novel insights for plastic biorecycling through protein engineering, which could lead to eco-friendly solutions to reduce the accumulation of plastic in landfills and natural environments.

Asunto(s)

Hidrolasas de Éster Carboxílico , Estabilidad de Enzimas , Especificidad por Sustrato , Cristalografía por Rayos X , Hidrolasas de Éster Carboxílico/metabolismo , Hidrolasas de Éster Carboxílico/química , Hidrolasas de Éster Carboxílico/genética , Ácidos Ftálicos/metabolismo , Ácidos Ftálicos/química , Ésteres/metabolismo , Ésteres/química , Modelos Moleculares , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Conformación Proteica , Concentración de Iones de Hidrógeno , Cinética , Hidrólisis , Dominio Catalítico , Temperatura