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1.
Adv Biochem Eng Biotechnol ; 170: 107-119, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-30847536

RESUMO

Aptazymes are synthetic molecules composed of an aptamer domain and a catalytic active nucleic acid unit, which may be a ribozyme or a DNAzyme. In these constructs the aptamer domain serves as a molecular switch that can regulate the catalytic activity of the ribozyme or DNAzyme subunit. This regulation is triggered by binding of the aptamers target molecule, which causes significant structural changes in the aptamer and thus in the entire aptazyme. Therefore, aptazymes function similar to allosteric enzymes, whose catalytic activity is regulated by binding of ligands (effectors) to allosteric sites due to alteration of the three-dimensional structure of the active site of the enzyme. In case of aptazymes, the allosteric site is composed of an aptamer. Aptazymes can be designed for different applications and have already been used in analytical assays as well as for the regulation of gene expression.


Assuntos
Aptâmeros de Nucleotídeos , DNA Catalítico , RNA Catalítico , Aptâmeros de Nucleotídeos/química , Catálise , DNA Catalítico/química , DNA Catalítico/metabolismo , Ligantes , Ligação Proteica , RNA Catalítico/química , RNA Catalítico/metabolismo
2.
Anticancer Res ; 39(10): 5531-5539, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31570447

RESUMO

BACKGROUND: Possible correlations between the expression of immune checkpoint molecules and prognosis in childhood acute leukemia were investigated. MATERIALS AND METHODS: The expression of programmed-death 1 (PD1), cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), and B- and T-lymphocyte attenuator (BTLA) was determined by flow cytometry on peripheral αß+ and γδ+ T-cells from patients with newly diagnosed acute lymphoblastic leukemia (ALL) (n=9) or acute myeloid leukemia (AML) (n=12), and from healthy volunteers (n=7). The expression of programmed-death ligand 1 (PD-L1), B7-1, B7-2, human leukocyte antigen-ABC (HLA-ABC), and herpesvirus-entry mediator (HVEM) ligands was determined on leukemia blasts. RESULTS: PD1 expression on αß+ and γδ+ T-cells was significantly higher in patients with ALL than in those with AML (p=0.0019 and 0.0239, respectively). CTLA-4 expression was moderately higher on αß+ and γδ+ T-cells in ALL (p=0.077 and 0.077, respectively), whereas HLA-ABC expression was significantly higher in AML blast cells (p=0.0182). The expression of CTLA-4 on γδ+ T-cells and the B7-2 ligand on blasts was higher in patients with high-risk ALL (p=0.02 and 0.02, respectively). In AML, PD1 expression on αß+ T-cells was higher in the intermediate-risk group (p=0.05), whereas HVEM expression was significantly higher in the low-risk group (p=0.02). Expression of CTLA-4 on γδ+ T-cells and PD-L1 on blasts were both associated with poor relapse-free survival outcomes in ALL (p=0.049). CONCLUSION: The higher expression of immune checkpoint molecules, in particular, CTLA-4 and PD-L1 are associated with a poorer prognosis in ALL, suggesting that selective use of the immune checkpoint blockade might improve the clinical outcomes in patients with ALL.


Assuntos
Leucemia/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Linfócitos T/imunologia , Doença Aguda , Adolescente , Adulto , Antígeno B7-H1/metabolismo , Antígeno CTLA-4/metabolismo , Criança , Pré-Escolar , Intervalo Livre de Doença , Feminino , Humanos , Lactente , Ligantes , Masculino , Pessoa de Meia-Idade , Recidiva , Adulto Jovem
3.
Cell Host Microbe ; 26(2): 160-162, 2019 08 14.
Artigo em Inglês | MEDLINE | ID: mdl-31415748

RESUMO

Host-microbiome interactions affect host physiology, but the underlying mechanisms are not well understood. Recent papers from Chen et al. (2019) and Colosimo et al. (2019) in this issue of Cell Host & Microbe demonstrate that metabolites produced by several members of the gut microbiota can efficiently activate host G protein-coupled receptors and influence host physiology.


Assuntos
Microbiota , Receptores Acoplados a Proteínas-G , Bactérias , Humanos , Ligantes , Transdução de Sinais
4.
Acta Crystallogr C Struct Chem ; 75(Pt 8): 1091-1101, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31380792

RESUMO

A new set of differently hydrated barium and strontium squarates, namely poly[[triaqua(µ-1,2-dioxocyclobut-3-ene-1,2-diolato)barium] monohydrate], {[Ba(C4O4)(H2O)3]·H2O}n (1), poly[[diaqua(µ-1,2-dioxocyclobut-3-ene-1,2-diolato)strontium] monohydrate], {[Sr(C4O4)(H2O)2]·H2O}n (2), and poly[[triaqua(µ-1,2-dioxocyclobut-3-ene-1,2-diolato)barium/strontium(0.85/0.15)] monohydrate], {[Ba0.85Sr0.15(C4O4)(H2O)3]·H2O}n (3), is reported. The study of their crystal structures indicates that all the complexes crystallize in the triclinic space group P-1. Complexes 1 and 3 have a rare combination of squarate units coordinated through monodentate O atoms to two different metal atoms and through two bidentate O atoms to three different metal atoms. Furthermore, they have three coordinated water molecules to give a coordination number of nine. The squarate ligands in complex 2 exhibit two different coordination modes: (i) monodentate O atoms coordinated to four different Sr atoms and (ii) two monodentate O atoms coordinated to two different metal atoms with the other two O atoms bidentate to four different Sr atoms. All the compounds decompose to give the respective carbonates when heated to 800 °C, as evidenced by thermogravimetry/differential thermal analysis (TG-DTA), which are clusters of nanoparticles. Complexes 1 and 3 show additional endothermic peaks at 811 and 820 °C, respectively, indicating the phase transition of BaCO3 from an orthorhombic (α-Pmcn) to a trigonal phase (ß-R3m). All three complexes have significant DNA-binding constants, ranging from 2.45 × 104 to 9.41 × 104 M-1 against EB-CT (ethidium bromide-calf thymus) DNA and protein binding constants ranging from 1.1 × 105 to 8.6 × 105 with bovine serum albumin. The in vitro cytotoxicity of the complexes is indicated by the IC50 values, which range from 128.8 to 261.3 µg ml-1. Complex 3 shows better BSA binding, antioxidant activity against the DPPH radical and cytotoxicity than complexes 1 and 2.


Assuntos
Antineoplásicos/farmacologia , Complexos de Coordenação/farmacologia , Ciclobutanos/farmacologia , Depuradores de Radicais Livres/farmacologia , Substâncias Intercalantes/farmacologia , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/metabolismo , Bário/química , Bovinos , Complexos de Coordenação/síntese química , Complexos de Coordenação/química , Complexos de Coordenação/metabolismo , Cristalografia por Raios X , Ciclobutanos/síntese química , Ciclobutanos/química , Ciclobutanos/metabolismo , DNA/metabolismo , Depuradores de Radicais Livres/síntese química , Depuradores de Radicais Livres/química , Depuradores de Radicais Livres/metabolismo , Humanos , Ligações de Hidrogênio , Substâncias Intercalantes/síntese química , Substâncias Intercalantes/química , Substâncias Intercalantes/metabolismo , Ligantes , Células MCF-7 , Estrutura Molecular , Ligação Proteica , Soroalbumina Bovina/metabolismo , Estrôncio/química , Água/química
5.
Chem Commun (Camb) ; 55(68): 10128-10131, 2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31386708

RESUMO

Fueled by the therapeutic potential of the epigenetic machinery, BET bromodomains have seen high interest as drug targets. Herein, we introduce different linkers to a BET bromodomain benzodiazepine ligand (I-BET762) to gauge its implications in the development of hybrid drugs, imaging probes and small molecule drug conjugates. Biophysical studies confirmed minimal disruption to binding of the BRD4 cavity by the synthesized entities, which includes imaging probes. Target engagement was confirmed in a cellular context, but poor membrane diffusion was found despite efficient localization in the nuclei after membrane disruption. Our study highlights challenges and opportunities for the successful design of benzodiazepine-derived drug-delivery systems.


Assuntos
Benzodiazepinas/farmacologia , Fluoresceínas/farmacologia , Corantes Fluorescentes/farmacologia , Proteínas Nucleares/antagonistas & inibidores , Benzodiazepinas/síntese química , Benzodiazepinas/química , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Desenho de Drogas , Fluoresceínas/síntese química , Fluoresceínas/química , Corantes Fluorescentes/síntese química , Corantes Fluorescentes/química , Humanos , Ligantes , Estrutura Molecular , Proteínas Nucleares/química , Domínios Proteicos
6.
J Chem Theory Comput ; 15(9): 5116-5134, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31386808

RESUMO

Enzymes play a pivotal role in all biological systems. These biomachines are the most effective catalysts known, dramatically enhancing the rate of reactions by more than 10 orders of magnitude relative to the uncatalyzed reactions in solution. Predicting the correct, mechanistically appropriate binding modes for substrate and product, as well as all reaction intermediates and transition states, along a reaction pathway is immensely challenging and remains an unsolved problem. In the present work, we developed an effective methodology for identifying probable binding modes of multiple ligand states along a reaction coordinate in an enzyme active site. The program is called EnzyDock and is a CHARMM-based multistate consensus docking program that includes a series of protocols to predict the chemically relevant orientation of substrate, reaction intermediates, transition states, product, and inhibitors. EnzyDock is based on simulated annealing molecular dynamics and Monte Carlo sampling and allows ligand, as well as enzyme side-chain and backbone flexibility. The program can employ many user-defined constraints and restraints and classical force field potentials, as well as a range of hybrid quantum mechanics-molecular mechanics potentials. Herein, we apply EnzyDock to several different kinds of problems. First, we study two terpene synthase reactions, namely bornyl diphosphate synthase and the bacterial diterpene synthase CotB2. Second, we use EnzyDock to predict reaction coordinate states in a pair of Diels-Alder reactions in the enzymes spirotetronate AbyU and LepI. Third, we study a couple of racemases: the cofactor-dependent serine racemase and the cofactor independent proline racemase. Finally, we study several cases of covalent docking involving the Michael addition reaction. For all systems we predict binding modes that are consistent with available experimental observations, as well as with theoretical modeling studies from the literature. EnzyDock provides a platform for generating mechanistic insight into enzyme reactions, useful and reliable starting points for in-depth multiscale modeling projects, and rational design of noncovalent and covalent enzyme inhibitors.


Assuntos
Racemases e Epimerases/química , Ligantes , Modelos Moleculares , Estrutura Molecular , Método de Monte Carlo , Engenharia de Proteínas , Teoria Quântica , Racemases e Epimerases/metabolismo
7.
J Chem Theory Comput ; 15(9): 4974-4981, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31402652

RESUMO

Predicting the costructure of small-molecule ligands and their respective target proteins has been a long-standing problem in drug discovery. For weak binding compounds typically identified in fragment-based screening (FBS) campaigns, determination of the correct binding site and correct binding mode is usually done experimentally via X-ray crystallography. For many targets of pharmaceutical interest, however, establishing an X-ray system which allows for sufficient throughput to support a drug discovery project is not possible. In this case, exploration of fragment hits becomes a very laborious and consequently slow process with the generation of protein/ligand cocrystal structures as the bottleneck of the entire process. In this work, we introduce a computational method which is able to reliably predict binding sites and binding modes of fragment-like small molecules using solely the structure of the apoprotein and the ligand's chemical structure as input information. The method is based on molecular dynamics simulations and Markov-state models and can be run as a fully automated protocol requiring minimal human intervention. We describe the application of the method to a representative subset of different target classes and fragments from historical FBS efforts at Boehringer Ingelheim and discuss its potential integration into the overall fragment-based drug discovery workflow.


Assuntos
Cadeias de Markov , Simulação de Dinâmica Molecular , Proteínas/química , Sítios de Ligação , Cristalografia por Raios X , Humanos , Ligantes
8.
Inorg Chem ; 58(16): 10778-10790, 2019 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-31386351

RESUMO

A new family of cyclometalated ruthenium(II) complexes [Ru(N^N)2(C^N)]+ derived from the π-extended benzo[h]imidazo[4,5-f]quinolone ligand appended with thienyl groups (n = 1-4, compounds 1-4) was prepared and its members were characterized for their chemical, photophysical, and photobiological properties. The lipophilicities of 1-4, determined as octanol-water partition coefficients (log Po/w), were positive and increased with the number of thienyl units. The absorption and emission bands of the C^N compounds were red-shifted by up to 200 nm relative to the analogous Ru(II) diimine systems. All of the complexes exhibited dual emission with the intraligand fluorescence (1IL, C^N-based) shifting to lower energies with increasing n and the metal-to-ligand charge transfer phosphorescence (3MLCT, N^N-based) remaining unchanged. Compounds 1-3 exhibited excited state absorption (ESA) profiles consistent with lowest-lying 3MLCT states when probed by nanosecond transient absorption (TA) spectroscopy with 532 nm excitation and had contributions from 1IL(C^N) states with 355 nm excitation. These assignments were supported by the lifetimes observed (<10 ns for the 1IL states and around 20 ns for the 3MLCT states) as well as a noticeable ESA for 3 with 355 nm excitation that did not occur with 532 nm excitation. Compound 4 was the only member of the family with two 3MLCT emissive lifetimes (15, 110 ns), and the TA spectra collected with both 355 and 532 nm excitation was assigned to the 3IL state, which was corroborated by its 4-6 µs lifetime. The ESA for 4 had a rise time of approximately 10 ns and an initial decay of 110 ns, which suggests a possible 3MLCT-3IL excited state equilibrium that results in delayed emission from the 3MLCT state. Compound 4 was nontoxic toward human skin melanoma cells (SKMEL28) in the dark (EC50 = >300 µM); 1-3 were cytotoxic and yielded EC50 values between 1 and 20 µM. The photocytotoxicites with visible light ranged from 87 nM with a phototherapeutic index (PI) of 13 for 1 to approximately 1 µM (PI = >267) for 4. With red light, EC50 values varied from 270 nM (PI = 21) for 3 to 12 µM for 4 (PI = >25). The larger PIs for 4, especially with visible light, were attributed to the much lower dark cytotoxicity for this compound. Because the dark cytotoxicity contributes substantially to the observed photocytotoxicity for 1-3, it was not possible to assess whether the 3IL state of 4 led to a much more potent phototoxic mechanism in the absence of dark toxicity. There was no stark contrast in cellular uptake and accumulation by laser scanning confocal and differential interference contrast microscopy to explain the large differences in dark toxicities between 1-3 and 4. Nevertheless, the study highlights a new family of Ru(II) C^N complexes where π-conjugation beyond a certain point results in low dark cytotoxicity with high photocytotoxicity, opposing the notion that cyclometalated Ru(II) systems are too toxic to be phototherapeutic agents.


Assuntos
Antineoplásicos/farmacologia , Complexos de Coordenação/farmacologia , Fármacos Fotossensibilizantes/farmacologia , Quinolonas/farmacologia , Rutênio/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Complexos de Coordenação/síntese química , Complexos de Coordenação/química , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Ligantes , Luz , Estrutura Molecular , Processos Fotoquímicos , Fotoquimioterapia , Fármacos Fotossensibilizantes/síntese química , Fármacos Fotossensibilizantes/química , Quinolonas/química , Rutênio/química
9.
Arch Virol ; 164(11): 2793-2797, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31440811

RESUMO

The DC-SIGN glycoprotein is responsible for the initial adhesion of dengue virus (DENV) to immune cells by the carbohydrate recognition domain (CRD). There are thirteen soluble and membrane-bound DC-SIGN isoforms, but the role of soluble isoforms in the DENV internalization process is not known. Five isoforms with an altered or absent CRD were identified, and three different soluble isoforms were used to confirm the interactions with mannose residues. The results show the loss of binding ability of one soluble isoform and binding ability of two of them. All of them will be used to verify their role in the DENV internalization process.


Assuntos
Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Vírus da Dengue/metabolismo , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Manose/metabolismo , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Ligação Viral , Internalização do Vírus , Sequência de Aminoácidos , Sequência de Bases , Dengue/virologia , Vírus da Dengue/genética , Ligantes , Ligação Proteica/genética , Isoformas de Proteínas/genética
10.
Phys Chem Chem Phys ; 21(33): 18149-18160, 2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31389436

RESUMO

Conformational entropies are of great interest when studying the binding of small ligands to proteins or the interaction of proteins. Unfortunately, there are no experimental methods available to measure conformational entropies of all groups in a protein. Instead, they are normally estimated from molecular dynamics (MD) simulations, although such methods show problems with convergence and correlation of motions, and depend on the accuracy of the underlying potential-energy function. Crystallographic atomic displacement parameters (also known as B-factors) are available in all crystal structures and contain information about the atomic fluctuations, which can be converted to entropies. We have studied whether B-factors can be employed to extract conformational entropies for proteins by comparing such entropies to those measured by NMR relaxation experiments or obtained from MD simulations in solution or in the crystal. Unfortunately, our results show that B-factor entropies are unreliable, because they include the movement and rotation of the entire protein, they exclude correlation of the movements and they include contributions other than the fluctuations, e.g. static disorder, as well as errors in the model and the scattering factors. We have tried to reduce the first problem by employing translation-libration-screw refinement, the second by employing a description of the correlated movement from MD simulations, and the third by studying only the change in entropy when a pair of ligands binds to the same protein, thoroughly re-refining the structures in exactly the same way and using the same set of alternative conformations. However, the experimental B-factors seem to be incompatible with fluctuations from MD simulations and the precision is too poor to give any reliable entropies.


Assuntos
Simulação de Dinâmica Molecular , Proteínas/química , Temperatura Ambiente , Cristalografia por Raios X , Entropia , Galectina 3/química , Ligantes , Muramidase/química , Conformação Proteica , Tripsina/química
11.
Int J Nanomedicine ; 14: 5017-5032, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31371944

RESUMO

Background: Epigallocatechin gallate (EGCG), the major anti-inflammatory compound in green tea, has been shown to suppress osteoclast (OC) differentiation. However, the low aqueous solubility of EGCG always leads to poor bioavailability, adverse effects, and several drawbacks for clinical applications. Purpose: In this study, we synthesized EGCG-capped gold nanoparticles (EGCG-GNPs) to solve the drawbacks for clinical uses of EGCG in bone destruction disorders by direct reduction of HAuCl4 in EGCG aqueous solution. Methods and Results: The obtained EGCG-GNPs were negatively charged and spherical. Theoretical calculation results suggested that EGCG was released from GNPs in an acidic environment. Cellular uptake study showed an obviously large amount of intracellular EGCG-GNPs without cytotoxicity. EGCG-GNPs exhibited better effects in reducing intracellular reactive oxygen species levels than free EGCG. A more dramatic anti-osteoclastogenic effect induced by EGCG-GNPs than free EGCG was observed in lipopolysaccharide (LPS)-stimulated bone marrow macrophages, including decreased formation of TRAP-positive multinuclear cells and actin rings. Meanwhile, EGCG-GNPs not only suppressed the mRNA expression of genetic markers of OC differentiation but also inhibited MAPK signaling pathways. Furthermore, we confirmed that EGCG-GNPs greatly reversed bone resorption in the LPS-induced calvarial bone erosion model in vivo, which was more effective than applying free EGCG, specifically in inhibiting the number of OCs, improving bone density, and preventing bone loss. Conclusion: EGCG-GNPs showed better anti-osteoclastogenic effect than free EGCG in vitro and in vivo, indicating their potential in anti-bone resorption treatment strategy.


Assuntos
Catequina/análogos & derivados , Ouro/farmacologia , Nanopartículas Metálicas/química , Osteogênese/efeitos dos fármacos , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/efeitos dos fármacos , Reabsorção Óssea/patologia , Catequina/farmacologia , Morte Celular/efeitos dos fármacos , Teoria da Densidade Funcional , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Liberação Controlada de Fármacos , Estabilidade de Medicamentos , Ligantes , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Nanopartículas Metálicas/ultraestrutura , Camundongos Endogâmicos BALB C , Modelos Biológicos , Ligante RANK/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Crânio/patologia , Transcrição Genética/efeitos dos fármacos
12.
Chem Pharm Bull (Tokyo) ; 67(7): 609-619, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31257315

RESUMO

To develop potent ligands for the vitamin D receptor (VDR), we designed and synthesized a series of vitamin D analogues with and without 22-alkyl substituents. These analogues exhibited agonistic, partial agonistic, or antagonistic activity. To elucidate the mechanism of action of the analogues, we conducted crystal structure analyses of the ligand-binding domain (LBD) of VDR complexed with the analogues. The VDR-LBD/agonist complex exhibited precise interactions, which clearly explained VDR agonism. The VDR-LBD/partial agonist complex showed two conformers (agonist and antagonist binding conformers) in a single crystal, demonstrating that partial agonism could be explained by the sum of the agonistic and antagonistic activities. Antagonist binding to the VDR-LBD structure was elucidated using both crystal structure analysis and in-solution structural analyses with the small-angle X-ray scattering (SAXS)-molecular dynamics (MD) and hydrogen/deuterium exchange coupled with mass spectrometry (HDX-MS) methods. Several antagonist-binding structures were detected. We found that the antagonist binding structures differed depending on the structure of the antagonist itself, and those structures clearly explained the VDR antagonism. Furthermore, the apo VDR-LBD structure without the ligand in the ligand-binding pocket was revealed and found to have an entrance to accommodate the ligand. Thus we elucidated the mechanisms of action of agonists, partial agonists, and antagonists based on structural changes (differences) in the receptor protein induced by ligand binding.


Assuntos
Ligantes , Receptores de Calcitriol/metabolismo , Sítios de Ligação , Cristalografia por Raios X , Humanos , Simulação de Dinâmica Molecular , Conformação Proteica em alfa-Hélice , Domínios Proteicos , Receptores de Calcitriol/agonistas , Receptores de Calcitriol/antagonistas & inibidores , Vitamina D/análogos & derivados , Vitamina D/metabolismo
13.
Inorg Chem ; 58(15): 10129-10138, 2019 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-31310108

RESUMO

A series of 16 "3 + 2" mixed-ligand complexes of the general composition [ReO(L1)(L2)] (H2L1a-H2L1d = tridentate thiosemicarbazones having a phenyl group with 4-H, 4-F, 3,5-di-F, and 4-CF3 substituents; HL2a-HL2d = bidentate N,N-diethyl-N'-benzoylthioureas with 4-H, 4-F, 3,5-di-F, and 4-CF3 substituents at the benzoyl groups) have been synthesized and characterized by spectroscopic methods and X-ray diffraction. Irrespective of the individual fluorine substitution, the complexes are stable and possess the same general structure. Some systematic electronic effects of the fluorine-substitution patterns of the ligands have been found on the 13C NMR chemical shifts of the N-C═N carbon atoms of the {L1}2- and the C═O carbon atoms of the {L2}- ligands. Antiparasitic properties of the rhenium complexes have been tested against epimastigotes and trypomastigotes forms of two Trypanosoma cruzi strains and the amastigotes form of one of them. The results of this study indicate that the activity of the rhenium complexes can clearly be modulated by fluorine substitution of their ligands. Some of the fluorinated compounds show a high activity against epimastigotes and trypomastigotes forms of the parasites. Reactions between (NBu4)[TcOCl4] and two representatives of the fluorinated ligands (H2L1b, 4-F-substituted, and H2L1c, 4-CF3-substituted) form stable complexes of the composition [TcOCl(L1b)] and [TcOCl(L1c)]. Subsequent reactions of these products with HL2b (4-F-substituted) give the corresponding [TcO(L1)(L2)] mixed-ligand complexes. Also, the technetium compounds are stable as solids and in solutions and have structures corresponding to those of their rhenium analogues.


Assuntos
Complexos de Coordenação/farmacologia , Halogenação , Rênio/farmacologia , Tiossemicarbazonas/farmacologia , Tioureia/farmacologia , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Complexos de Coordenação/síntese química , Complexos de Coordenação/química , Cristalografia por Raios X , Haplorrinos , Ligantes , Modelos Moleculares , Estrutura Molecular , Testes de Sensibilidade Parasitária , Rênio/química , Tiossemicarbazonas/química , Tioureia/química , Tripanossomicidas/síntese química , Tripanossomicidas/química
14.
Gene ; 712: 143945, 2019 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-31279712

RESUMO

Reactive oxygen species, generated in all the aerobic organisms, can cause oxidative stress. Excessive ROS may become a source of carcinogen due to DNA damage, lipid peroxidation, cell injury, and cell death. In order to prevent these adverse effects of ROS, antioxidant enzymes have evolved in aerobic organisms. Catalase is a major antioxidant enzyme that breaks down excessive H2O2 and inhibits apoptotic cell death. Here we molecularly characterized catalase from red-lip mullet. The cDNA sequence of LhCAT consists of an ORF of 1545 bp, which encodes a 527 amino acid peptide (~60 kDa). Based on bioinformatics analysis, LhCAT possesses a domain architecture characteristic of catalases, including a catalase proximal active site signature and a catalase proximal heme-ligand signature. It also has heme and NADPH binding sites homologous to previously described catalases. Pairwise alignment with its homologs revealed that LhCAT shares 95.1% identity with Oplegnathus fasciatus catalase and 97.4% similarity with Sparus aurata catalase. An uprooted phylogenetic tree demonstrated that LhCAT resides in a clade with catalases from other teleosts and exhibits a close relationship with Oplegnathus fasciatus catalase. Among twelve tissue types, we observed the highest LhCAT mRNA expression in the liver, followed by blood. Immune challenge by Lactococcus garvieae, or Poly I:C in the blood or spleen resulted in up-regulation at 24 h post injection. We also tested the antioxidant activity of recombinant LhCAT against hydrogen peroxide and found its optimal concentration to be 12.5 µg/mL. Collectively, these data suggested that LhCAT play an important role in antioxidant defense and immune response of red-lip mullet.


Assuntos
Catalase/metabolismo , Proteínas de Peixes/metabolismo , Smegmamorpha , Adjuvantes Imunológicos , Animais , Antioxidantes/metabolismo , Catalase/genética , DNA Complementar/genética , Proteínas de Peixes/genética , Perfilação da Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Heme/química , Peróxido de Hidrogênio/química , Sistema Imunitário , Ligantes , Fígado/enzimologia , Estresse Oxidativo , Filogenia , RNA Mensageiro/metabolismo , Espécies Reativas de Oxigênio , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Regulação para Cima
15.
Nat Commun ; 10(1): 2915, 2019 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-31266946

RESUMO

The bile acid-sensing transcription factor farnesoid X receptor (FXR) regulates multiple metabolic processes. Modulation of FXR is desired to overcome several metabolic pathologies but pharmacological administration of full FXR agonists has been plagued by mechanism-based side effects. We have developed a modulator that partially activates FXR in vitro and in mice. Here we report the elucidation of the molecular mechanism that drives partial FXR activation by crystallography- and NMR-based structural biology. Natural and synthetic FXR agonists stabilize formation of an extended helix α11 and the α11-α12 loop upon binding. This strengthens a network of hydrogen bonds, repositions helix α12 and enables co-activator recruitment. Partial agonism in contrast is conferred by a kink in helix α11 that destabilizes the α11-α12 loop, a critical determinant for helix α12 orientation. Thereby, the synthetic partial agonist induces conformational states, capable of recruiting both co-repressors and co-activators leading to an equilibrium of co-activator and co-repressor binding.


Assuntos
Receptores Citoplasmáticos e Nucleares/agonistas , Receptores Citoplasmáticos e Nucleares/química , Animais , Linhagem Celular , Proteínas Correpressoras/genética , Proteínas Correpressoras/metabolismo , Humanos , Ligações de Hidrogênio , Ligantes , Fígado/metabolismo , Espectroscopia de Ressonância Magnética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ligação Proteica , Conformação Proteica em alfa-Hélice , Receptores Citoplasmáticos e Nucleares/genética , Receptores Citoplasmáticos e Nucleares/metabolismo
16.
Chem Commun (Camb) ; 55(65): 9590-9605, 2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31334709

RESUMO

The study of artificial receptor molecules with the intention to mimic enzyme-substrate binding processes and catalysis in nature has always been a traditional area of research in supramolecular chemistry. Along this line, our group has developed a family of porphyrin cage compounds based on glycoluril and employed these in host-guest binding studies, as components of allosterically controlled self-assembled processes, in which structural changes in the cage upon complexation of a guest or a ligand change binding equilibria, and as enzyme mimics in supramolecular catalysis. In a recently started research program aimed at developing a new molecular approach to long-term data storage, porphyrin cage compounds are studied as molecular machines to encode information into synthetic polymer chains. In this Feature Article we will give an overview of the above aspects of our porphyrin cage compounds and place them in the context of related systems reported in the literature.


Assuntos
Alquinos/química , Materiais Biomiméticos/química , Imidazóis/química , Porfirinas/química , Receptores Artificiais/química , Materiais Biomiméticos/síntese química , Catálise , Imidazóis/síntese química , Ligantes , Metaloporfirinas/síntese química , Metaloporfirinas/química , Nanotecnologia/métodos , Porfirinas/síntese química , Receptores Artificiais/síntese química , Estereoisomerismo , Termodinâmica
17.
J Agric Food Chem ; 67(33): 9286-9294, 2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31339733

RESUMO

Natural aryl hydrocarbon (AHR) ligands have been identified in food and herbal medicines, and they may exhibit beneficial activity in humans. In this study, white button (WB) feeding significantly decreased AHR target gene expression in the small intestine of both conventional and germ-free mice. High-performance liquid chromatography (HPLC) fractionation and ultra-high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) combined with an AHR-responsive cell-based luciferase gene reporter assay were used to isolate and characterize benzothiazole (BT) derivatives and 6-methylisoquinoline (6-MIQ) as AHR modulators from WB mushrooms. The study showed dose-dependent changes of AHR transformation determined by the cell-based luciferase gene reporter assay and transcription of CYP1A1 in human Caco-2 cells by BT derivatives and 6-MIQ. These findings suggested that WB mushroom contains new classes of natural AHR modulators and demonstrated HPLC fractionation and UHPLC-MS/MS combined with a cell-based luciferase gene reporter assay as a useful approach for isolation and characterization of the previously unidentifed AHR modulators from natural products.


Assuntos
Agaricus/química , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Receptores de Hidrocarboneto Arílico/genética , Animais , Benzotiazóis/química , Benzotiazóis/isolamento & purificação , Benzotiazóis/farmacologia , Linhagem Celular , Cromatografia Líquida de Alta Pressão , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A1/metabolismo , Genes Reporter , Humanos , Isoquinolinas/química , Isoquinolinas/isolamento & purificação , Isoquinolinas/farmacologia , Ligantes , Camundongos , Extratos Vegetais/farmacologia , Receptores de Hidrocarboneto Arílico/metabolismo , Espectrometria de Massas em Tandem , Ativação Transcricional/efeitos dos fármacos , Verduras/química
18.
Zhongguo Yi Xue Ke Xue Yuan Xue Bao ; 41(3): 300-306, 2019 Jun 30.
Artigo em Chinês | MEDLINE | ID: mdl-31282322

RESUMO

Objective To investigate the effects of different inflammatory factors on hepatocyte kinase receptor(Eph)and ligand(ephrin)in human periodontal ligament fibroblasts(hPDLFs).Methods hPDLFs were stimulated with either 10 ng/ml tumor necrosis factor-α(TNF-α)or 10 ng/ml interleukin(IL)-1ß,and then the expressions of Eph and ephrin at both mRNA and protein levels were determined at 0,1,2,6,12,and 24 hours.Results The levels of Eph receptors and ephrin ligand changed in a time-dependent manner in human periodontal ligament fibroblasts after treatment with TNF-α or IL-1ß. The expression of ephrinA2 significantly increased in both groups within 24 hours(all P<0.05). In the TNF-α group,the mRNA expression of ephrinA2 significantly increased at 1 h and was significant higher that in the IL-1ß group at 24 h(P<0.05). EphB4 showed a time-dependent decline after a short period of high expression.Conclusions Both TNF-α and IL-1ß can cause changes in the expressions of Eph receptors and ephrin ligands in hPDLFs. The changes induced by both are consistent,although the effect of TNF-α is more pronounced.


Assuntos
Efrinas/metabolismo , Fibroblastos , Ligamento Periodontal/citologia , Receptores da Família Eph/metabolismo , Células Cultivadas , Humanos , Interleucina-1beta/farmacologia , Ligantes , Fator de Necrose Tumoral alfa/farmacologia
19.
J Enzyme Inhib Med Chem ; 34(1): 1373-1379, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31347933

RESUMO

Butyrylcholinesterase (BChE) plays an important role in the progression of the Alzheimer's disease. In this study, we used a structure-based virtual screening (VS) approach to discover new BChE inhibitors. A ligand database was filtered and docked to the BChE protein using Glide program. The outcome from VS was filtered and the top ranked hits were thoroughly examined for their fitting into the protein active site. Consequently, the best 38 hits were selected for in vitro testing using Ellman's method, and six of which showed inhibition activity for BChE. Interestingly, the most potent hit (Compound 4) exhibited inhibitory activity against the BChE enzyme in the low micromolar level with an IC50 value of 8.3 µM. Hits obtained from this work can act as a starting point for future SAR studies to discover new BChE inhibitors as anti-Alzheimer agents.


Assuntos
Butirilcolinesterase/metabolismo , Inibidores da Colinesterase/síntese química , Inibidores da Colinesterase/farmacologia , Descoberta de Drogas/métodos , Simulação de Acoplamento Molecular , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/enzimologia , Inibidores da Colinesterase/química , Humanos , Ligantes , Estrutura Molecular , Relação Estrutura-Atividade
20.
J Phys Chem A ; 123(28): 5995-6002, 2019 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-31268326

RESUMO

High-resolution X-ray crystallography and two-dimensional NMR studies demonstrate that water-mediated conventional hydrogen-bonding interactions (N-H···N, O-H···N, etc.) bridging two or more amino acid residues contribute to the stability of proteins and protein-ligand complexes. In this work, we have investigated single water-mediated selenium hydrogen-bonding interactions (unconventional hydrogen-bonding) between amino acid residues in proteins through extensive protein data bank (PDB) analysis coupled with gas-phase spectroscopy and quantum chemical calculation of a model complex consisting of indole, dimethyl selenide, and water. Here, indole and dimethyl selenide represent the amino acid residues tryptophan and selenomethionine, respectively. The current investigation demonstrates that the most stable structure of the model complex observed in the IR spectroscopy mimics single water-mediated selenium hydrogen-bonded structural motifs present in the crystal structures of proteins. The present work establishes that water-mediated Se hydrogen-bonding interactions are ubiquitous in proteins and the number of these interactions observed in the PDB is more than that of direct Se hydrogen-bonds present there.


Assuntos
Proteínas/química , Selênio/química , Água/química , Biologia Computacional , Cristalografia por Raios X , Bases de Dados de Proteínas , Ligações de Hidrogênio , Indóis/química , Ligantes , Modelos Moleculares , Compostos Organosselênicos/química , Teoria Quântica , Selenometionina/química , Espectrofotometria Infravermelho , Triptofano/química
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