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1.
Molecules ; 29(19)2024 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-39407546

RESUMO

This study characterized the binding mechanisms of the lectin cMoL (from Moringa oleifera seeds) to carbohydrates using spectroscopy and molecular dynamics (MD). The interaction with carbohydrates was studied by evaluating lectin fluorescence emission after titration with glucose or galactose (2.0-11 mM). The Stern-Volmer constant (Ksv), binding constant (Ka), Gibbs free energy (∆G), and Hill coefficient were calculated. After the urea-induced denaturation of cMoL, evaluations were performed using fluorescence spectroscopy, circular dichroism (CD), and hemagglutinating activity (HA) evaluations. The MD simulations were performed using the Amber 20 package. The decrease in Ksv revealed that cMoL interacts with carbohydrates via a static mechanism. The cMoL bound carbohydrates spontaneously (ΔG < 0) and presented a Ka on the order of 102, with high selectivity for glucose. Protein-ligand complexes were stabilized by hydrogen bonds and hydrophobic interactions. The Hill parameter (h~2) indicated that the binding occurs through the cMoL dimer. The loss of HA at urea concentrations at which the fluorescence and CD spectra indicated protein monomerization confirmed these results. The MD simulations revealed that glucose bound to the large cavity formed between the monomers. In conclusion, the biotechnological application of cMoL lectin requires specific methods or media to improve its dimeric protein structure.


Assuntos
Simulação de Dinâmica Molecular , Moringa oleifera , Ligação Proteica , Sementes , Moringa oleifera/química , Sementes/química , Lectinas de Plantas/química , Multimerização Proteica , Carboidratos/química , Dicroísmo Circular , Lectinas/química , Lectinas/metabolismo , Espectrometria de Fluorescência , Conformação Proteica , Termodinâmica , Ligação de Hidrogênio
2.
Spectrochim Acta A Mol Biomol Spectrosc ; 314: 124176, 2024 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-38513314

RESUMO

Here, we presented a second-order scattering sensor based on the Zn0.97La0.03O compound (LaZnO) for selective and stable detection of glycated albumin (GA, glycemic long-term biomarker). The LaZnO sample was obtained through the co-precipitation method and then characterized using microscopic and spectroscopic techniques. Furthermore, the selectivity, molecular interference, temporal stability, and pH effects of the LaZnO SOS signal in the absence and presence of GA were investigated. The results indicate the stability of the SOS signal over more than 60 days. Assays conducted within the pH range of 5 to 8 indicate that the detection of GA remains unaffected under the given conditions. Selectivity studies show that the SOS signal of LaZnO is reduced only upon contact with GA, while interference studies show that detection is not affected by other chemical species. Additionally, the calibration curve test showed high sensitivity of the material, with a detection limit of 0.55 µg/ml. All the results suggest that LaZnO can deliver efficiency, selectivity, accuracy, and fast response as a GA biosensor, emphasizing LaZnO's usefulness in detecting protein biomarkers.


Assuntos
Albumina Sérica Glicada , Produtos Finais de Glicação Avançada , Albumina Sérica/metabolismo , Biomarcadores , Zinco , Glicemia
3.
Mol Pharm ; 20(8): 4021-4030, 2023 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-37382244

RESUMO

The ability to bind plasma proteins helps in comprehending relevant aspects related to the pharmacological properties of many drugs. Despite the vital role of the drug mubritinib (MUB) in the prophylaxis of various diseases, its interaction with carrier proteins still needs to be clarified. The present work focuses on the interaction between MUB and Human serum albumin (HSA), investigated by employing multispectroscopic, biochemical, and molecular docking approaches. The results reveal that MUB has quenched HSA intrinsic fluorescence (following a static mechanism) by attaching very close (r = 6.76 Å) and with moderate affinity (Kb ≈ 104 M-1) to the protein site I (mainly by H-bonds, hydrophobic and Van der Waals forces). On one side, the HSA-MUB interaction has been accompanied by a slight disturbance in the HSA chemical environment (around the Trp residue) and protein secondary structure modifications. On another side, MUB competitively inhibits HSA esterase-like activity, which is very similar to other Tyrosine kinase inhibitors, and evidence that protein functional alterations have been triggered by MUB interaction. In summary, all of the presented observations can shed light on diverse pharmacological factors associated with drug administration.


Assuntos
Albumina Sérica Humana , Humanos , Albumina Sérica Humana/química , Simulação de Acoplamento Molecular , Sítios de Ligação , Ligação Proteica , Transporte de Elétrons , Espectrometria de Fluorescência , Termodinâmica , Dicroísmo Circular
4.
ACS Chem Neurosci ; 12(24): 4500-4511, 2021 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-34808043

RESUMO

Tyrosine kinase inhibitors (TKIs) are antitumor compounds that prevent the phosphorylation of proteins in a biological environment. However, the multitarget performance of TKIs promotes them as possible candidates for drug repositioning. In this work, interaction and inhibition studies through spectroscopic and computational techniques to evaluate the binding effectiveness of lapatinib and pazopanib TKIs to acetylcholinesterase (AChE) are reported. The results indicated potent inhibition at the µM level. The types of inhibition were identified, with pazopanib acting through non-competitive inhibition and lapatinib through acompetitive inhibition. The fluorescence suppression studies indicate a static mechanism for lapatinib-AChE and pazopanib-AChE systems, with a binding constant in the order of 105 M-1. The obtained thermodynamic parameters reveal interactions driven by van der Waals forces and hydrogen bonds in the lapatinib-AChE system (ΔH° and ΔS° < 0). In contrast, the pazopanib-AChE system shows positive ΔH° and ΔS°, characteristic of hydrophobic interactions. The Foster resonance energy transfer study supports the fluorescence studies performed. The 3D fluorescence studies suggest changes in the microenvironment of the tryptophan and tyrosine residues of the protein in contact with lapatinib and pazopanib. The results suggest effective inhibition and moderate interaction of the drugs with AChE, making them interesting for conducting more in-depth repositioning studies as AChE inhibitors.


Assuntos
Acetilcolinesterase , Preparações Farmacêuticas , Acetilcolinesterase/metabolismo , Sítios de Ligação , Indazóis , Lapatinib , Ligação Proteica , Pirimidinas/farmacologia , Sulfonamidas , Termodinâmica
5.
Colloids Surf B Biointerfaces ; 207: 112006, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34343910

RESUMO

Small organic molecules have been extensively applied to achieve enzymatic inhibition. Although numerous efforts have been made to deliver efficient inhibitors, small inhibitors applications are hindered by many drawbacks. Moreover, reporters comprising nanoparticle inhibitory activity against enzymes are very scarce in the literature. In this scenario, carbon nanodots (CDs) emerge as promising candidates for efficient enzyme inhibition due to their unique properties. Here, CDs specific molecular characteristics (core composition and chemical surface groups) have been investigated to produce a more potent enzyme inhibition. Mushroom tyrosinase (mTyr) has been adopted as an enzymatic prototype. The CDs revealed a high affinity to mTyr (Ka ≈ 106 M-1), mainly through hydrophobic forces and followed by slight mTyr structural alteration. CDs competitively inhibit mTyr, with low inhibition constant (KI = 517.7 ±â€¯17.0 nM), which is up 70 fold smaller then the commercial inhibitor (kojic acid) and the starch nanoparticles previously reported. The results expose that the CDs act as a hydrophobic agglomerate with carboxyl groups on its surface, mimicking characteristics found on small molecule inhibitors (but with superior performance). All these results highlight the CD excellent potential as an efficient low toxic Tyr inhibitor, opening the prospect of using these nanoparticles in the cosmetic and food industries.


Assuntos
Carbono , Nanopartículas , Inibidores Enzimáticos/farmacologia , Monofenol Mono-Oxigenase , Amido
6.
Int J Biol Macromol ; 168: 676-685, 2021 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-33220373

RESUMO

Egletes viscosa is a plant with therapeutic value due to its antibacterial, antinociceptive and gastroprotective properties. This study aimed to purify, characterize, and evaluate the cytotoxicity of a lectin (EgviL) from the floral capitula of E. viscosa. The lectin was isolated from saline extract through precipitation with ammonium sulfate followed by Sephadex G-75 chromatography. The molecular mass and isoelectric point (pI) of EgviL were determined as well as its temperature and pH stability. Physical-chemical parameters of interaction between EgviL and carbohydrates were investigated by fluorescence quenching and 1H nuclear magnetic resonance (NMR). Cytotoxicity was investigated against human peripheral blood mononuclear cells (PBMCs) and neoplastic cells. EgviL (28.8 kDa, pI 5.4) showed hemagglutinating activity stable towards heating until 60 °C and at the pH range 5.0-7.0. This lectin is able to interact through hydrophobic and electrostatic bonds with galactose and glucose, respectively. EgviL reduced the viability of PBMCs only at the highest concentration tested (100 µg/mL) while was toxic to Jurkat E6-1 cells with IC50 of 24.1 µg/mL,inducing apoptosis. In summary, EgviL is a galactose/glucose-binding protein with acidic character, stable to heating and with cytotoxic effect on leukemic cells.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Asteraceae/química , Leucócitos Mononucleares/citologia , Lectinas de Plantas/farmacologia , Antineoplásicos Fitogênicos/química , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Precipitação Química , Estabilidade de Medicamentos , Galactose/metabolismo , Glucose/metabolismo , Testes de Hemaglutinação , Humanos , Interações Hidrofóbicas e Hidrofílicas , Concentração Inibidora 50 , Ponto Isoelétrico , Células Jurkat , Leucócitos Mononucleares/efeitos dos fármacos , Células MCF-7 , Lectinas de Plantas/química
7.
Int J Biol Macromol ; 136: 1034-1041, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31233796

RESUMO

Many skin disorders and diseases are related to tyrosinase activity, in particular, due to the vital role played by this enzyme in the melanogenic process. Although numerous natural and synthetic tyrosinase inhibitors have been published, substantial efforts have been made to understand the influence of tyrosinase inhibition on the viability of melanoma cells. Here, we assess the impact of two keto-derivatives: 2-acetyl-furan (F1), furfural-acetone (F2), and two carboxyl-derivatives: 2-furan-acrylic acid (F3), 5-methyl-2-furan-acrylic acid (F4), on the mushroom tyrosinase (mTYR) activity, by applying spectroscopic, kinetic and theoretical techniques. From an exploratory and theoretical point of view, results indicated that albeit all furans bind tightly to and inhibit mTYR very efficient, carboxyl-furan derivatives presented best inhibitory activities than keto- derivatives and performed the inhibition competitively and reversible. Moreover, we examined the influence of carboxyl derivative on the viability of melanoma cells. Results expose differential toxicity of these furan derivatives, which indicates a piece of evidence that furan inhibition activity may be related to its toxicity against B16F10 cells.


Assuntos
Antineoplásicos/farmacologia , Inibidores Enzimáticos/farmacologia , Furanos/farmacologia , Melanoma/patologia , Monofenol Mono-Oxigenase/antagonistas & inibidores , Agaricales/enzimologia , Animais , Antineoplásicos/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Inibidores Enzimáticos/metabolismo , Furanos/metabolismo , Humanos , Camundongos , Simulação de Acoplamento Molecular , Monofenol Mono-Oxigenase/química , Monofenol Mono-Oxigenase/metabolismo , Conformação Proteica
8.
Int J Biol Macromol ; 122: 289-297, 2019 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-30401647

RESUMO

Here, we evaluate spiroacridines as inhibitors of tyrosinase, a key enzyme to melanogenesis. For this purpose, the spiroacridines 3-(acridin-9-yl)-N-benzylidene-2-cyanoacrylohydrazide (AMTAC-01) and 3-(acridin-9-yl)-2-cyano-N-(4-metoxybenzylidene)-acrylohydrazide (AMTAC-02) were synthesized and their enzymatic inhibition types and mechanisms were investigated. In addition, the interaction of these compounds with the enzyme were studied by UV-Vis spectroscopy, spectrofluorimetry, 1H NMR titration as well as molecular docking. Spectroscopic results reveals that the acridine derivatives interact strongly (Ka ≅ 104 - 105 M-1) with the mushroom tyrosinase and the enzyme undergoes small structural modifications due to the interaction with AMTAC-01 compound. The interaction studies support the enzymatic inhibition results, which suggests that AMTAC-01 compounds inhibit the enzyme reversibly and follows a noncompetitive type (AMTAC-01) and mixed type (AMTAC-02) of inhibition. Nevertheless, AMTAC-02 (IC50 = 96.29 µM) inhibits the enzyme more effectively than AMTAC-01 (IC50 = 189.40 µM), which suggests a highly relevant role of AMTAC-02's methoxy group to the inhibition activity, which is confirmed by docking studies to mushroom tyrosinase. Docking also indicates this interaction to be absent in human tyrosinase. SIGNIFICANCE: Based on previous results which evidenced the relevant activity of two spiroacridinic compounds for cell growth inhibition against melanoma cells, here we improve our understanding about the spiroacridines in the biological media by exploring the molecular mechanism that govern the activities of these two compounds using mushroom tyrosinase (mTYR) enzyme as molecular target. The paper not only will have a major impact upon molecular mechanism that regulates melanin inhibition by spiroacridinic compounds, but also by guiding the search for enzyme inhibitors and the development of new anti-melanoma prophylaxis.


Assuntos
Acridinas/química , Acridinas/farmacologia , Simulação de Acoplamento Molecular , Monofenol Mono-Oxigenase/antagonistas & inibidores , Monofenol Mono-Oxigenase/metabolismo , Compostos de Espiro/química , Acridinas/metabolismo , Inibidores Enzimáticos/química , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Humanos , Ligantes , Monofenol Mono-Oxigenase/química , Ligação Proteica , Conformação Proteica
9.
Int J Biol Macromol ; 92: 467-475, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27435006

RESUMO

Two new spiro-acridines were synthesized by introducing cyano-N-acylhydrazone between the acridine and phenyl rings followed by spontaneous cyclization. The final compounds (E)-1'-(benzylideneamino)-5'-oxo-1',5'-dihydro-10H-spiro[acridine-9,2'-pyrrole]-4'-carbonitrile (AMTAC-01) and (E)-1'-((4-methoxybenzylidene)amino)-5'-oxo-1',5'-dihydro-10H-spiro[acridine-9,2'-pyrrole]-4'-carbonitrile (AMTAC-02) were evaluated for their interactions with calf thymus DNA, antiproliferative and human topoisomerase I and IIα inhibitory activities. Both compounds presented ability to bind DNA. The binding constant determined by UV-vis spectroscopy was found to be 104M-1. Antiproliferative assay demonstrated that AMTAC-01 and AMTAC-02 were most active against prostate and melanoma tumor cell lines, respectively. The compound did not present Topo I inhibitory activity. However, both derivatives displayed topoisomerase IIα inhibitory activity comparable to amsacrine, and AMTAC-02 was more potent than AMTAC-01 with methoxy substituent group on phenyl ring. This study demonstrates that the new derivatives are promising molecules with topoisomerase IIα inhibitory and antiproliferative activities.


Assuntos
Acridinas/farmacologia , DNA Topoisomerases/metabolismo , DNA/metabolismo , Compostos de Espiro/farmacologia , Inibidores da Topoisomerase/farmacologia , Acridinas/síntese química , Acridinas/química , Animais , Espectroscopia de Ressonância Magnética Nuclear de Carbono-13 , Bovinos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Humanos , Espectroscopia de Prótons por Ressonância Magnética , Espectrometria de Fluorescência , Espectrofotometria Ultravioleta , Compostos de Espiro/síntese química , Compostos de Espiro/química
10.
Eur J Med Chem ; 104: 148-56, 2015 Nov 02.
Artigo em Inglês | MEDLINE | ID: mdl-26454648

RESUMO

A series of thiophene-2-thiosemicarbazones derivatives (5-14) was synthesized, characterized and evaluated for their antitumor activity. They were tested in vitro against human tumor cell lines through the colorimetric method. The results revealed that compounds 7 and 9 were the most effective in inhibiting 50% of the cell growth after 48 h of treatment. As compound 7 showed a potent antiproliferative profile, it has been chosen for further studies in 786-0 cell line by flow cytometry. Treatments with compound 7 (50 µM) induced early phosphatidylserine exposure after 18 h of exposure and this process progressed phosphatidylserine exposure with loss of cell membrane integrity after 24 h of treatment, suggesting a time-dependent cell death process. Regarding the cell cycle profile, no changes were observed after treatment with compound 7 (25 µM), suggesting a mechanism of cell death independent on the cell cycle. The in vivo studies show that compound 7 possess low acute toxicity, being the doses of 30-300 mgKg(-1) chosen for studies in Ehrlich solid tumor model in mice. All doses were able to inhibit tumor development being the lowest one the most effective. Our findings highlight thiophene-2-thiosemicarbazones as a promising class of compounds for further studies concerning new anticancer therapies.


Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Carcinoma de Ehrlich/tratamento farmacológico , Tiofenos/farmacologia , Tiossemicarbazonas/farmacologia , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Carcinoma de Ehrlich/patologia , Ciclo Celular/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Estrutura Molecular , Relação Estrutura-Atividade , Tiofenos/administração & dosagem , Tiofenos/síntese química , Tiofenos/química , Tiossemicarbazonas/administração & dosagem , Tiossemicarbazonas/síntese química , Tiossemicarbazonas/química
11.
Proteins ; 78(16): 3386-95, 2010 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-20848643

RESUMO

Many plant pathogenic bacteria rely on effector proteins to suppress defense and manipulate host cell mechanisms to cause disease. The effector protein PthA modulates the host transcriptome to promote citrus canker. PthA possesses unusual protein architecture with an internal region encompassing variable numbers of near-identical tandem repeats of 34 amino acids termed the repeat domain. This domain mediates protein-protein and protein-DNA interactions, and two polymorphic residues in each repeat unit determine DNA specificity. To gain insights into how the repeat domain promotes protein-protein and protein-DNA contacts, we have solved the structure of a peptide corresponding to 1.5 units of the PthA repeat domain by nuclear magnetic resonance (NMR) and carried out small-angle X-ray scattering (SAXS) and spectroscopic studies on the entire 15.5-repeat domain of PthA2 (RD2). Consistent with secondary structure predictions and circular dichroism data, the NMR structure of the 1.5-repeat peptide reveals three α-helices connected by two turns that fold into a tetratricopeptide repeat (TPR)-like domain. The NMR structure corroborates the theoretical TPR superhelix predicted for RD2, which is also in agreement with the elongated shape of RD2 determined by SAXS. Furthermore, RD2 undergoes conformational changes in a pH-dependent manner and upon DNA interaction, and shows sequence similarities to pentatricopeptide repeat (PPR), a nucleic acid-binding motif structurally related to TPR. The results point to a model in which the RD2 structure changes its compactness as it embraces the DNA with the polymorphic diresidues facing the interior of the superhelix oriented toward the nucleotide bases.


Assuntos
Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , DNA Bacteriano/metabolismo , Sequências Repetitivas de Aminoácidos , Motivos de Aminoácidos , Sequência de Aminoácidos , Dicroísmo Circular , Concentração de Íons de Hidrogênio , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Dados de Sequência Molecular , Peptídeos/química , Peptídeos/metabolismo , Estrutura Terciária de Proteína
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