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1.
Braz. j. biol ; 83: e247604, 2023. tab, graf
Article in English | MEDLINE, LILACS, VETINDEX | ID: biblio-1339370

ABSTRACT

Abstract In the current report, we studied the possible inhibitors of COVID-19 from bioactive constituents of Centaurea jacea using a threefold approach consisting of quantum chemical, molecular docking and molecular dynamic techniques. Centaurea jacea is a perennial herb often used in folk medicines of dermatological complaints and fever. Moreover, anticancer, antioxidant, antibacterial and antiviral properties of its bioactive compounds are also reported. The Mpro (Main proteases) was docked with different compounds of Centaurea jacea through molecular docking. All the studied compounds including apigenin, axillarin, Centaureidin, Cirsiliol, Eupatorin and Isokaempferide, show suitable binding affinities to the binding site of SARS-CoV-2 main protease with their binding energies -6.7 kcal/mol, -7.4 kcal/mol, -7.0 kcal/mol, -5.8 kcal/mol, -6.2 kcal/mol and -6.8 kcal/mol, respectively. Among all studied compounds, axillarin was found to have maximum inhibitor efficiency followed by Centaureidin, Isokaempferide, Apigenin, Eupatorin and Cirsiliol. Our results suggested that axillarin binds with the most crucial catalytic residues CYS145 and HIS41 of the Mpro, moreover axillarin shows 5 hydrogen bond interactions and 5 hydrophobic interactions with various residues of Mpro. Furthermore, the molecular dynamic calculations over 60 ns (6×106 femtosecond) time scale also shown significant insights into the binding effects of axillarin with Mpro of SARS-CoV-2 by imitating protein like aqueous environment. From molecular dynamic calculations, the RMSD and RMSF computations indicate the stability and dynamics of the best docked complex in aqueous environment. The ADME properties and toxicity prediction analysis of axillarin also recommended it as safe drug candidate. Further, in vivo and in vitro investigations are essential to ensure the anti SARS-CoV-2 activity of all bioactive compounds particularly axillarin to encourage preventive use of Centaurea jacea against COVID-19 infections.


Resumo No presente relatório, estudamos os possíveis inibidores de Covid-19 de constituintes bioativos de Centaurea jacea usando uma abordagem tripla que consiste em técnicas de química quântica, docking molecular e dinâmica molecular. Centaurea jacea é uma erva perene frequentemente usada em remédios populares de doenças dermatológicas e febre. Além disso, as propriedades anticâncer, antioxidante, antibacteriana e antiviral de seus compostos bioativos também são relatadas. A Mpro (proteases principais) foi acoplada a diferentes compostos de Centaurea jacea por meio de docking molecular. Todos os compostos estudados, incluindo apigenina, axilarina, Centaureidina, Cirsiliol, Eupatorina e Isokaempferide, mostram afinidades de ligação adequadas ao sítio de ligação da protease principal SARS-CoV-2 com suas energias de ligação -6,7 kcal / mol, -7,4 kcal / mol, - 7,0 kcal / mol, -5,8 kcal / mol, -6,2 kcal / mol e -6,8 kcal / mol, respectivamente. Dentre todos os compostos estudados, a axilarina apresentou eficiência máxima de inibidor, seguida pela Centaureidina, Isokaempferida, Apigenina, Eupatorina e Cirsiliol. Nossos resultados sugeriram que a axilarina se liga aos resíduos catalíticos mais cruciais CYS145 e HIS41 do Mpro, além disso a axilarina mostra 5 interações de ligações de hidrogênio e 5 interações hidrofóbicas com vários resíduos de Mpro. Além disso, os cálculos de dinâmica molecular em uma escala de tempo de 60 ns (6 × 106 femtossegundos) também mostraram percepções significativas sobre os efeitos de ligação da axilarina com Mpro de SARS-CoV-2 por imitação de proteínas como o ambiente aquoso. A partir de cálculos de dinâmica molecular, os cálculos RMSD e RMSF indicam a estabilidade e dinâmica do melhor complexo ancorado em ambiente aquoso. As propriedades ADME e a análise de previsão de toxicidade da axilarina também a recomendaram como um candidato a medicamento seguro. Além disso, as investigações in vivo e in vitro são essenciais para garantir a atividade anti-SARS-CoV-2 de todos os compostos bioativos, particularmente a axilarina, para encorajar o uso preventivo de Centaurea jacea contra infecções por Covid-19.


Subject(s)
Humans , Pharmaceutical Preparations , Centaurea , COVID-19 , Protease Inhibitors , Molecular Dynamics Simulation , Molecular Docking Simulation , SARS-CoV-2
2.
Protein & Cell ; (12): 877-888, 2021.
Article in English | WPRIM | ID: wpr-922482

ABSTRACT

A new coronavirus (SARS-CoV-2) has been identified as the etiologic agent for the COVID-19 outbreak. Currently, effective treatment options remain very limited for this disease; therefore, there is an urgent need to identify new anti-COVID-19 agents. In this study, we screened over 6,000 compounds that included approved drugs, drug candidates in clinical trials, and pharmacologically active compounds to identify leads that target the SARS-CoV-2 papain-like protease (PLpro). Together with main protease (M


Subject(s)
Antiviral Agents/therapeutic use , Binding Sites , COVID-19/virology , Coronavirus Papain-Like Proteases/metabolism , Crystallography, X-Ray , Drug Evaluation, Preclinical , Drug Repositioning , High-Throughput Screening Assays/methods , Humans , Imidazoles/therapeutic use , Inhibitory Concentration 50 , Molecular Dynamics Simulation , Mutagenesis, Site-Directed , Naphthoquinones/therapeutic use , Protease Inhibitors/therapeutic use , Protein Structure, Tertiary , Recombinant Proteins/isolation & purification , SARS-CoV-2/isolation & purification
3.
Braz. J. Pharm. Sci. (Online) ; 56: e17420, 2020. tab, graf
Article in English | LILACS | ID: biblio-1142490

ABSTRACT

Dengue fever has emerged as a big threat to human health since the last decade owing to high morbidity with considerable mortalities. The proposed study aims at the in silico investigation of the inhibitory action against DENV4-NS1 of phytochemicals from two local medicinal plants of Pakistan. Non-Structural Protein 1 of Dengue Virus 4 (DENV4-NS1) is known to be involved in the replication and maturation of viron in the host cells. A total of 129 phytochemicals (50 from Tanacetum parthenium and 79 from Silybum marianum) were selected for this study. The tertiary structure of DENV4-NS1 was predicted based on homology modelling using Modeller 9.18 and the structural stability was evaluated using molecular dynamics simulations. Absorption, distribution, metabolism, excretion and toxicity (ADMET) along with the drug-likeness was also predicted for these phytochemicals using SwissADME and PreADMET servers. The results of ADMET and drug-likeness predictions exhibited that 54 phytochemicals i.e. 25 from Tanacetum parthenium and 29 from Silybum marianum showed effective druglikeness. These phytochemicals were docked against DENV4-NS1 using AutoDock Vina and 18 most suitable phytochemicals with binding affinities ≤ -6.0 kcal/mol were selected as potential inhibitors for DENV4-NS1. Proposed study also exploits the novel inhibitory action of Jaceidin, Centaureidin, Artecanin, Secotanaparthenolide, Artematin, Schizolaenone B, Isopomiferin, 6, 8-Diprenyleriodictyol, and Anthraxin against dengue virus. It is concluded that the screened 18 phytochemicals have strong inhibition potential against Dengue Virus 4.


Subject(s)
Computer Simulation , Proteins/classification , Dengue , Dengue Virus , Phytochemicals/analysis , Plants, Medicinal/metabolism , Pharmacokinetics , Tanacetum parthenium/adverse effects , Molecular Dynamics Simulation
4.
Chinese Journal of Biotechnology ; (12): 1819-1828, 2019.
Article in Chinese | WPRIM | ID: wpr-771750

ABSTRACT

We review major computational chemistry techniques applied in industrial enzyme studies, especially approaches intended for guiding enzyme engineering. These include molecular mechanics force field and molecular dynamics simulation, quantum mechanical and combined quantum mechanical/molecular mechanical approaches, electrostatic continuum models, molecular docking, etc. These approaches are essentially introduced from the following two angles for viewing: one is about the methods themselves, including the basic concepts, the primary computational results, and potential advantages and limitations; the other is about obtaining valuable information from the respective calculations to guide the design of mutants and mutant libraries.


Subject(s)
Enzymes , Chemistry , Genetics , Metabolism , Molecular Docking Simulation , Molecular Dynamics Simulation , Mutant Proteins , Chemistry , Genetics , Metabolism , Protein Engineering , Quantum Theory , Static Electricity
5.
Chinese Journal of Biotechnology ; (12): 636-646, 2019.
Article in Chinese | WPRIM | ID: wpr-771345

ABSTRACT

Glutamate decarboxylase, a unique pyridoxal 5'-phosphate-dependent enzyme, catalyzes α-decarboxylation of L-glutamate to γ-aminobutyrate. However, glutamate decarboxylase from different sources has the common problem of poor thermostability that affects its application in industry. In this study, proline was introduced at 13 different positions in glutamate decarboxylase by using the design strategy of homologous sequence alignment between Thermococcus kodakarensis and Lactobacillus brevis CGMCC No.1306. A mutant enzyme G364P with higher thermostability was obtained. Compared to the wild type, thermostability of the mutant G364P was significantly improved, the half-life time (t1/2) at 55 °C and the semi-inactivation temperature (T₅₀ ¹⁵) of the mutant G364P increased 19.4 min and 5.3 °C, respectively, while kcat/Km of the mutant enzyme remained nearly unchanged. Further analysis of their thermostability by molecular dynamics simulations were performed. The root mean square deviation of G364P and root mean square fluctuation in the loop region including G364 were lower than the wild type at 313 K for 10 ns, and G364P increased one hydrophobic interaction in the loop region. It proves that mutation of flexible 364-Gly to rigid proline endows glutamate decarboxylase with enhanced thermostability.


Subject(s)
Glutamate Decarboxylase , Glutamic Acid , Lactobacillus brevis , Molecular Dynamics Simulation , Proline
6.
Electron. j. biotechnol ; 31: 93-99, Jan. 2018. ilus, graf, tab
Article in English | LILACS | ID: biblio-1022150

ABSTRACT

Background: Peptidoglycan (PGN) recognition proteins (PGRPs) are important pattern recognition receptors of the host innate immune system that are involved in the immune defense against bacterial pathogens. PGRPs have been characterized in several fish species. The PGN-binding ability is important for the function of PGRPs. However, the PGRP-PGN interaction mechanism in fish remains unclear. In the present study, the 3-D model of a long PGRP of half-smooth tongue sole (Cynoglossus semilaevis) (csPGRP-L), a marine teleost with great economic value, was constructed through the comparative modeling method, and the key amino acids involved in the interaction with Lys-type PGNs and Dap-type PGNs were analyzed by molecular dynamics and molecular docking methods. Results: csPGRP-L possessed a typical PGRP structure, consisting of five ß-sheets and four α-helices. Molecular docking showed that the van der Waals forces had a slightly larger contribution than Coulombic interaction in the csPGRP-L-PGN complex. Moreover, the binding energies of csPGRP-L-PGNs computed by MM-PBSA method revealed that csPGRP-L might selectively bind both types of MTP-PGNs and MPP-PGNs. In addition, the binding energy of each residue of csPGRP-L was also calculated, revealing that the residues involved in the interaction with Lys-type PGNs were different from that with Dap-type PGNs. Conclusions: The 3-D structure of csPGRP-L possessed typical PGRP structure and might selectively bind both types of MTP- and MPP-PGNs, which provided useful insights to understanding the functions of fish PGRPs.


Subject(s)
Animals , Tongue/immunology , Flatfishes/immunology , Flatfishes/metabolism , Binding Sites , Flatfishes/genetics , Peptidoglycan , Carrier Proteins , Toll-Like Receptors , Molecular Dynamics Simulation , Molecular Docking Simulation , Ligands
7.
Article in English | WPRIM | ID: wpr-772963

ABSTRACT

Androgen receptor (AR) is a ligand-activated transcription factor that plays a pivotal role in the development and progression of many severe diseases such as prostate cancer, muscle atrophy, and osteoporosis. Binding of ligands to AR triggers the conformational changes in AR that may affect the recruitment of coactivators and downstream response of AR signaling pathway. Therefore, AR ligands have great potential to treat these diseases. In this study, we searched for novel AR ligands by performing a docking-based virtual screening (VS) on the basis of the crystal structure of the AR ligand binding domain (LBD) in complex with its agonist. A total of 58 structurally diverse compounds were selected and subjected to LBD affinity assay, with five of them (HBP1-3, HBP1-17, HBP1-38, HBP1-51, and HBP1-58) exhibiting strong binding to AR-LBD. The IC values of HBP1-51 and HBP1-58 are 3.96 µM and 4.92 µM, respectively, which are even lower than that of enzalutamide (Enz, IC = 13.87 µM), a marketed second-generation AR antagonist. Further bioactivity assays suggest that HBP1-51 is an AR agonist, whereas HBP1-58 is an AR antagonist. In addition, molecular dynamics (MD) simulations and principal components analysis (PCA) were carried out to reveal the binding principle of the newly-identified AR ligands toward AR. Our modeling results indicate that the conformational changes of helix 12 induced by the bindings of antagonist and agonist are visibly different. In summary, the current study provides a highly efficient way to discover novel AR ligands, which could serve as the starting point for development of new therapeutics for AR-related diseases.


Subject(s)
Androgen Receptor Antagonists , Pharmacology , Androgens , Metabolism , Pharmacology , Biological Assay , Cell Line, Tumor , Drug Discovery , Methods , Humans , Ligands , Male , Molecular Docking Simulation , Molecular Dynamics Simulation , Phenylthiohydantoin , Pharmacology , Principal Component Analysis , Prostatic Neoplasms , Drug Therapy , Protein Binding , Physiology , Protein Conformation , Receptors, Androgen , Metabolism
8.
São Paulo; s.n; s.n; 2018. 85 p. tab, graf, ilus.
Thesis in Portuguese | LILACS | ID: biblio-982084

ABSTRACT

A doença de Chagas, causada pelo parasita Trypanosoma cruzi, acomete entre 6 a 8 milhões de pessoas em todo o mundo. Conhecida como tripanossomíase americana, por ter sido considerada endêmica apenas na América Latina, esta doença, se espalhou para outros continentes devido aos movimentos migratórios se tornando um problema de sáude mundial. Estima-se que 56.000 novos casos e cerca de 12.000 mortes por complicações relacionadas à doença de Chagas anualmente. A quimioterapia disponível para o tratamento é composta apenas por dois fármacos, nifurtimox e benznidazol, no entanto são pouco eficazes na fase crônica da doença. Estes fármacos apresentarem, ainda, efeitos adversos graves e resistência por parte de algumas cepas do parasita. Diante deste panorama, é iminente a necessidade da busca de novos fármacos contra T. cruzi. Para a busca racional de novos quimiterapicos antiparasitários é fundamental a identificação e caracterização de vias metabólicas essenciais à sobrevivência dos parasitas. Assim, a enzima sirtuína 2 - Silent Information Regulator 2 (Sir2), tem importante papel para a infecção por T. cruzi, pois está totalmente envolvida no seu ciclo celular do parasita. Esta é uma enzima NAD+ dependente da classe III histona desacetilases, e se mostra como um interessante alvo bioquímico para o desenvolvimento de antichagásicos. A disponibilidade do sequenciamento genômico da Sir2 nos permite utilizar estratégias de planejamento de fármaco baseado no receptor (SBDD - Structure Based Drug Design) na identificação de candidatos a fármacos para essa doença. Entre as técnicas modernas de SBDD utilizadas, a triagem virtual possibilita identificar e selecionar inibidores enzimáticos potentes e seletivos para o alvo escolhido. Assim, neste trabalho, foi construído por meio da técnica de modelagem comparativa o modelo da enzima Sir2 de T. cruzi. Uma simulação por dinâmica molecular de 200ns, foi realizada para averiguar a estabilidade do modelo obtido. Diante da estabilização do modelo a partir de 100ns, o mesmo foi validado utilizando análise de clusters, RMSD (Root-mean-square Deviation) e análises de frequência de ligações de hidrogênio com o Cofator (NAD+) e os aminoácidos do sítio de catálise foram observadas, estes passos de simulação e validação foram realizados no programa DESMOND. Com o modelo robusto, os campos de interações moleculares (MIFs) foram gerados no programa GRID (Molecular Discovery v2.1) com o intuito de elucidar as regiões favoráveis a interação com a enzima em relação a propriedades físico-químicas da Sir2. A partir dos MIFs favoráveis a Sir2 de T. cruzi foi possível a construção de dois modelos farmacofóricos, o qual se baseou nas interações do Cofator (NAD+) e o sítio de catálise (Nicotinamida). O mesmo foi apliacdo como filtro para Triagem Virtual no programa UNITY da plataforma SYBYL X 2.0, utilizando os bancos de dados ZINC15 e GSK. A triagem resultou na seleção de 8 compostos candidatos a inibidores. Destes foram adquiridos 6 compostos por serem considerados mais promissores devido a complementariedade molecular. Estes foram testados contra a enzima de T. cruzi Sri2. Após o ensaio foi possível avaliar a potência de 4 compostos, sendo o composto CDMS-01 (IC50 = 39,9uM) o mais promissor que será submetido à processos de otimização molecular


Chagas disease, caused by the parasite Trypanosoma cruzi, affects between 6 and 8 million people worldwide. Also known as American trypanosomiasis, because it is considered endemic only in Latin America, but has spread to other continents due to migratory movements. It is estimated that 56,000 new cases and about 12,000 deaths from complications related to Chagas disease annually. The chemotherapy available for treatment consists of only two drugs, nifurtimox and benznidazole, however these are poorly effective in the chronic phase. These drugs also have serious adverse effects and resistance from strains of the parasite. Faced with this scenario, the need to search for new drugs against T. cruzi is imminent. For the drug planning for new antiparasitic chemotherapics, the identification and characterization of metabolic pathways essential to the survival of parasites is fundamental. Therewith, the sirtuin 2 - Silent Information Regulator 2 (Sir2) enzyme has an important role for T. cruzi infection, since Sir2 in the parasite is totally involved in its cell cycle. This is an NAD+-dependent enzyme of class III histone deacetylases, and it shows an interesting biochemical target for the development of antichagasic. The availability of Sir2 genomic sequencing allows us to use SBDD (Structure Based Drug Design) strategies in identifying drug candidates for this disease. Among the modern techniques of SBDD used, virtual screening makes it possible to identify and select potent and selective enzyme inhibitors for the chosen target. The model of the T. cruzi Sir2 enzyme was constructed using the comparative modeling technique. A molecular dynamics simulation of 200ns was performed to ascertain the stability of the obtained model. Considering the stabilization of the model from 100ns, it was validated using cluster analysis, Root-mean-square Deviation (RMSD) and hydrogen bond frequency analyzes with Cofator (NAD+) and the amino acids of the catalysis site were observed, these simulation and validation steps were performed in the DESMOND program. With the robust model, the molecular interaction fields (MIFs) were generated in the GRID program (Molecular Discovery v2.1) in order to elucidate the regions favorable to the interaction with the enzyme in relation to the physicalchemical properties of Sir2. From the MIFs favorable to Sir2 of T. cruzi it was possible to construct two pharmacophoric models, which was based on the interactions of Cofator (NAD+) and the catalysis site (Nicotinamide). It was also applied as a Virtual screening filter in the UNITY program of the SYBYL X 2.0 platform, using the ZINC15 and GSK databases. Screening resulted in the selection of 8 inhibitor candidate compounds. Six compounds were obtained from the screening, because they were considered more promising, and were tested against T. cruzi Sri2 enzyme. After the assay it was possible to evaluate the potency of 4 compounds, the most promising compound being CDMS-01 (IC50 = 39.9 µM) that will be submitted to molecular optimization processes


Subject(s)
Trypanosoma cruzi/pathogenicity , Sirtuin 2/analysis , Validation Study , Drug Compounding , Sirtuin 2/antagonists & inhibitors , Molecular Dynamics Simulation , Antiparasitic Agents
9.
Genomics & Informatics ; : 142-146, 2017.
Article in English | WPRIM | ID: wpr-192018

ABSTRACT

More effective production of human insulin is important, because insulin is the main medication that is used to treat multiple types of diabetes and because many people are suffering from diabetes. The current system of insulin production is based on recombinant DNA technology, and the expression vector is composed of a preproinsulin sequence that is a fused form of an artificial leader peptide and the native proinsulin. It has been reported that the sequence of the leader peptide affects the production of insulin. To analyze how the leader peptide affects the maturation of insulin structurally, we adapted several in silico simulations using 13 artificial proinsulin sequences. Three-dimensional structures of models were predicted and compared. Although their sequences had few differences, the predicted structures were somewhat different. The structures were refined by molecular dynamics simulation, and the energy of each model was estimated. Then, protein-protein docking between the models and trypsin was carried out to compare how efficiently the protease could access the cleavage sites of the proinsulin models. The results showed some concordance with experimental results that have been reported; so, we expect our analysis will be used to predict the optimized sequence of artificial proinsulin for more effective production.


Subject(s)
Computer Simulation , DNA, Recombinant , Humans , Insulin , Molecular Dynamics Simulation , Proinsulin , Protein Sorting Signals , Trypsin
10.
Immune Network ; : 237-249, 2017.
Article in English | WPRIM | ID: wpr-22201

ABSTRACT

Using biomarkers as prediction tools or therapeutic targets can be a valuable strategy in transplantation. Recent studies identified biomarkers of acute rejection (AR) and operational tolerance (TOL) through the application of meta-analysis. In this study, we comparatively analyzed the signature genes in acute rejection and operational tolerance seen in human allogeneic transplantations using massive bioinformatical meta-analysis. To identify the signature genes in opposite immunological conditions, AR and TOL, we first collected the 1,252 gene expression data specifically intended for those circumstances. Then we excluded based on biological cut-values, Principal Component Analysis (PCA) as well as Multi-Dimensional Scaling (MDS). Using differentially expressed genes (DEGs) from meta-analysis, we then applied a ranked scoring system to identify the signature genes of AR and TOL. We identified 53 up-regulated and 32 down-regulated signature genes in acute rejection condition. Among them, ISG20, CXCL9, CXCL10, CCL19, FCER1G, PMSE1, UBD are highly expressed in AR condition. In operational tolerance, we identified 110 up-regulated and 48 down-regulated signature genes. TCL1A, BLNK, MS4A1, EBF1, IGHM are up-regulated in TOL condition. These genes are highly representative of AR or TOL across the different organs such as liver, kidney and heart. Since immune response is the sum of complex biological and molecular dynamics, these signature genes as well as pathway analysis using a systems biology approach could be used to catch the insights of the certain pathways that would be overlooked with the conventional gene-level comparative analysis.


Subject(s)
Biomarkers , Gene Expression , Graft Rejection , Heart , Humans , Kidney , Liver , Molecular Dynamics Simulation , Principal Component Analysis , Systems Biology , Transplantation Tolerance
11.
Mem. Inst. Oswaldo Cruz ; 111(12): 721-730, Dec. 2016. tab, graf
Article in English | LILACS | ID: biblio-829257

ABSTRACT

The main challenge in the control of malaria has been the emergence of drug-resistant parasites. The presence of drug-resistant Plasmodium sp. has raised the need for new antimalarial drugs. Molecular modelling techniques have been used as tools to develop new drugs. In this study, we employed virtual screening of a pyrazol derivative (Tx001) against four malaria targets: plasmepsin-IV, plasmepsin-II, falcipain-II, and PfATP6. The receiver operating characteristic curves and area under the curve (AUC) were established for each molecular target. The AUC values obtained for plasmepsin-IV, plasmepsin-II, and falcipain-II were 0.64, 0.92, and 0.94, respectively. All docking simulations were carried out using AutoDock Vina software. The ligand Tx001 exhibited a better interaction with PfATP6 than with the reference compound (-12.2 versus -6.8 Kcal/mol). The Tx001-PfATP6 complex was submitted to molecular dynamics simulations in vacuum implemented on an NAMD program. The ligand Tx001 docked at the same binding site as thapsigargin, which is a natural inhibitor of PfATP6. Compound TX001 was evaluated in vitro with a P. falciparum strain (W2) and a human cell line (WI-26VA4). Tx001 was discovered to be active against P. falciparum (IC50 = 8.2 µM) and inactive against WI-26VA4 (IC50 > 200 µM). Further ligand optimisation cycles generated new prospects for docking and biological assays.


Subject(s)
Humans , Antimalarials/chemistry , Aspartic Acid Endopeptidases/chemistry , Cysteine Endopeptidases/chemistry , Molecular Dynamics Simulation , Protozoan Proteins/chemistry , Thapsigargin/chemistry , Computational Biology/methods , Molecular Targeted Therapy/methods
12.
Protein & Cell ; (12): 501-515, 2016.
Article in English | WPRIM | ID: wpr-757411

ABSTRACT

β/γ-Crystallins are predominant structural proteins in the cytoplasm of lens fiber cells and share a similar fold composing of four Greek-key motifs divided into two domains. Numerous cataract-causing mutations have been identified in various β/γ-crystallins, but the mechanisms underlying cataract caused by most mutations remains uncharacterized. The S228P mutation in βB1-crystallin has been linked to autosomal dominant congenital nuclear cataract. Here we found that the S228P mutant was prone to aggregate and degrade in both of the human and E. coli cells. The intracellular S228P aggregates could be redissolved by lanosterol. The S228P mutation modified the refolding pathway of βB1-crystallin by affecting the formation of the dimeric intermediate but not the monomeric intermediate. Compared with native βB1-crystallin, the refolded S228P protein had less packed structures, unquenched Trp fluorophores and increased hydrophobic exposure. The refolded S228P protein was prone to aggregate at the physiological temperature and decreased the protective effect of βB1-crystallin on βA3-crystallin. Molecular dynamic simulation studies indicated that the mutation decreased the subunit binding energy and modified the distribution of surface electrostatic potentials. More importantly, the mutation separated two interacting loops in the C-terminal domain, which shielded the hydrophobic core from solvent in native βB1-crystallin. These two interacting loops are highly conserved in both of the N- and C-terminal domains of all β/γ-crystallins. We propose that these two interacting loops play an important role in the folding and structural stability of β/γ-crystallin domains by protecting the hydrophobic core from solvent access.


Subject(s)
Amino Acid Substitution , Cataract , Genetics , Metabolism , HeLa Cells , Humans , Molecular Dynamics Simulation , Mutation, Missense , Protein Aggregation, Pathological , Genetics , Metabolism , Protein Domains , Protein Structure, Secondary , Proteolysis , beta-Crystallin B Chain , Chemistry , Genetics , Metabolism
13.
Article in English | WPRIM | ID: wpr-177270

ABSTRACT

The vitamin D receptor (VDR) is a member of the nuclear receptor (NR) superfamily. The VDR binds to active vitamin D3 metabolites, which stimulates downstream transduction signaling involved in various physiological activities such as calcium homeostasis, bone mineralization, and cell differentiation. Quercetin is a widely distributed flavonoid in nature that is known to enhance transactivation of VDR target genes. However, the detailed molecular mechanism underlying VDR activation by quercetin is not well understood. We first demonstrated the interaction between quercetin and the VDR at the molecular level by using fluorescence quenching and saturation transfer difference (STD) NMR experiments. The dissociation constant (K(d)) of quercetin and the VDR was 21.15 ± 4.31 µM, and the mapping of quercetin subsites for VDR binding was performed using STD-NMR. The binding mode of quercetin was investigated by a docking study combined with molecular dynamics (MD) simulation. Quercetin might serve as a scaffold for the development of VDR modulators with selective biological activities.


Subject(s)
Calcification, Physiologic , Calcium , Cell Differentiation , Cholecalciferol , Fluorescence , Homeostasis , Molecular Dynamics Simulation , Quercetin , Receptors, Calcitriol , Transcriptional Activation , Vitamin D , Vitamins
14.
Chinese Journal of Biotechnology ; (12): 669-682, 2016.
Article in Chinese | WPRIM | ID: wpr-337432

ABSTRACT

Faldaprevir analogue molecule (FAM) has been reported to effectively inhibit the catalytic activity of HCV NS3/4A protease, making it a potential lead compound against HCV. A series of HCV NS3/4A protease crystal structures were analyzed by bioinformatics methods, and the FAM-HCV NS3/4A protease crystal structure was chosen for this study. A 20.4 ns molecular dynamics simulation of the complex consists of HCV NS3/4A protease and FAM was conducted. The key amino acid residues for interaction and the binding driving force for the molecular recognition between the protease and FAM were identified from the hydrogen bonds and binding free energy analyses. With the driving force of hydrogen bonds and van der Waals, FAM specifically bind to the active pocket of HCV NS3/4A protease, including V130-S137, F152-D166, D77-D79 and V55, which agreed with the experimental data. The effect of R155K, D168E/V and V170T site-directed mutagenesis on FAM molecular recognition was analyzed for their effect on drug resistance, which provided the possible molecular explanation of FAM resistance. Finally, the system conformational change was explored by using free energy landscape and conformational cluster. The result showed four kinds of dominant conformation, which provides theoretical basis for subsequent design of Faldaprevir analogue inhibitors based on the structure of HCV NS3/4A protease.


Subject(s)
Antiviral Agents , Chemistry , Carrier Proteins , Chemistry , Drug Resistance, Viral , Endopeptidases , Hepacivirus , Molecular Dynamics Simulation , Mutagenesis, Site-Directed , Oligopeptides , Chemistry , Protease Inhibitors , Chemistry , Serine Proteases , Thiazoles , Chemistry , Viral Nonstructural Proteins , Chemistry
15.
Chinese Medical Journal ; (24): 860-867, 2016.
Article in English | WPRIM | ID: wpr-328143

ABSTRACT

<p><b>BACKGROUND</b>Congenital cataract (CC) is the leading cause of visual impairment or blindness in children worldwide. Because of highly genetic and clinical heterogeneity, a molecular diagnosis of the lens disease remains a challenge.</p><p><b>METHODS</b>In this study, we tested a three-generation Chinese family with autosomal dominant CCs by targeted sequencing of 45 CC genes on next generation sequencing and evaluated the pathogenicity of the detected mutation by protein structure, pedigree validation, and molecular dynamics (MD) simulation.</p><p><b>RESULTS</b>A novel 15 bp deletion on GJA8 (c.426_440delGCTGGAGGGGACCCT or p. 143_147delLEGTL) was detected in the family. The deletion, concerned with an in-frame deletion of 5 amino acid residues in a highly evolutionarily conserved region within the cytoplasmic loop domain of the gap junction channel protein connexin 50 (Cx50), was in full cosegregation with the cataract phenotypes in the family but not found in 1100 control exomes. MD simulation revealed that the introduction of the deletion destabilized the Cx50 gap junction channel, indicating the deletion as a dominant-negative mutation.</p><p><b>CONCLUSIONS</b>The above results support the pathogenic role of the 15 bp deletion on GJA8 in the Chinese family and demonstrate targeted genes sequencing as a resolution to molecular diagnosis of CCs.</p>


Subject(s)
Adolescent , Adult , Aged , Cataract , Genetics , Connexins , Chemistry , Genetics , Female , Gene Deletion , High-Throughput Nucleotide Sequencing , Humans , Male , Middle Aged , Molecular Dynamics Simulation , Mutation
16.
Genomics & Informatics ; : 53-61, 2016.
Article in English | WPRIM | ID: wpr-213649

ABSTRACT

Toxoplasma gondii is an intracellular Apicomplexan parasite and a causative agent of toxoplasmosis in human. It causes encephalitis, uveitis, chorioretinitis, and congenital infection. T. gondii invades the host cell by forming a moving junction (MJ) complex. This complex formation is initiated by intermolecular interactions between the two secretory parasitic proteins—namely, apical membrane antigen 1 (AMA1) and rhoptry neck protein 2 (RON2) and is critically essential for the host invasion process. By this study, we propose two potential leads, NSC95522 and NSC179676 that can efficiently target the AMA1 hydrophobic cleft, which is a hotspot for targeting MJ complex formation. The proposed leads are the result of an exhaustive conformational search-based virtual screen with multilevel precision scoring of the docking affinities. These two compounds surpassed all the precision levels of docking and also the stringent post docking and cumulative molecular dynamics evaluations. Moreover, the backbone flexibility of hotspot residues in the hydrophobic cleft, which has been previously reported to be essential for accommodative binding of RON2 to AMA1, was also highly perturbed by these compounds. Furthermore, binding free energy calculations of these two compounds also revealed a significant affinity to AMA1. Machine learning approaches also predicted these two compounds to possess more relevant activities. Hence, these two leads, NSC95522 and NSC179676, may prove to be potential inhibitors targeting AMA1-RON2 complex formation towards combating toxoplasmosis.


Subject(s)
Chorioretinitis , Drug Design , Encephalitis , Humans , Hydrophobic and Hydrophilic Interactions , Machine Learning , Membranes , Molecular Docking Simulation , Molecular Dynamics Simulation , Neck , Parasites , Pliability , Toxoplasma , Toxoplasmosis , Uveitis
17.
Protein & Cell ; (12): 403-416, 2016.
Article in English | WPRIM | ID: wpr-757127

ABSTRACT

YfiBNR is a recently identified bis-(3'-5')-cyclic dimeric GMP (c-di-GMP) signaling system in opportunistic pathogens. It is a key regulator of biofilm formation, which is correlated with prolonged persistence of infection and antibiotic drug resistance. In response to cell stress, YfiB in the outer membrane can sequester the periplasmic protein YfiR, releasing its inhibition of YfiN on the inner membrane and thus provoking the diguanylate cyclase activity of YfiN to induce c-di-GMP production. However, the detailed regulatory mechanism remains elusive. Here, we report the crystal structures of YfiB alone and of an active mutant YfiB(L43P) complexed with YfiR with 2:2 stoichiometry. Structural analyses revealed that in contrast to the compact conformation of the dimeric YfiB alone, YfiB(L43P) adopts a stretched conformation allowing activated YfiB to penetrate the peptidoglycan (PG) layer and access YfiR. YfiB(L43P) shows a more compact PG-binding pocket and much higher PG binding affinity than wild-type YfiB, suggesting a tight correlation between PG binding and YfiB activation. In addition, our crystallographic analyses revealed that YfiR binds Vitamin B6 (VB6) or L-Trp at a YfiB-binding site and that both VB6 and L-Trp are able to reduce YfiB(L43P)-induced biofilm formation. Based on the structural and biochemical data, we propose an updated regulatory model of the YfiBNR system.


Subject(s)
Amino Acid Sequence , Bacterial Proteins , Chemistry , Genetics , Metabolism , Binding Sites , Biofilms , Crystallography, X-Ray , Cyclic GMP , Metabolism , Dimerization , Molecular Dynamics Simulation , Molecular Sequence Data , Mutagenesis , Protein Structure, Quaternary , Pseudomonas aeruginosa , Metabolism , Sequence Alignment , Tryptophan , Chemistry , Metabolism , Vitamin B 6 , Chemistry , Metabolism
18.
Rev. cuba. inform. méd ; 8(supl.1)2016.
Article in Spanish | LILACS, CUMED | ID: biblio-844908

ABSTRACT

Se presenta un método para la detección de semejanza entre moléculas basado en macheo inexacto de grafos. Se parte del grafo molecular completo ponderado en sus vértices por propiedades químico-físicas particionadas sobre los mismos, se reduce el grafo por el procedimiento CALEDE que define Centros Descriptores o fragmentos de primer orden, los cuales son subgrafos ponderados por la suma de los valores de los vértices ponderados individualmente a su vez, y se construyen fragmentos denominados de segundo orden que incluyen la distancia entre los centros de masas de ambos centros descriptores. Se presenta el método de búsqueda aplicado a una base de datos de más de 300 moléculas con sus respectivas estructuras en tres dimensiones. Esos compuestos se encuentran evaluados como anticancerígenos en la base de datos de compuestos del NCBI-USA. En el experimento computacional se encuentra que, en dependencia de la función de similitud empleada, es posible detectar compuestos que a pesar de poseer diferente topología, poseen valores de las propiedades empleadas para el macheo lo cual sugiere la presencia de potenciales farmacóforos como hallazgo relevante, lo cual constituiría un novedoso enfoque para el diseño computacional de fármacos(AU)


A method for detecting similarity between molecules based on inexact matching graph is presented. We start from a complete molecular graph vertices weighted by several hybrid indices. The molecular graph is reduced by CALEDE procedure, which define descriptors centers or first order fragments. These fragments are subgraphs weighted with the sum of values of the vertices weighted with the hybrid indices. It also define second order fragments by including the distance between the centers of mass of both descriptors centers. The search method applied to a database of over 300 molecules with their respective threedimensional structures is presented. These compounds are reported in the NCBI-USA database of compounds whish were evaluated in anticancer tests. In the computational experiment, depending on the similarity function used, is possible to detect compounds that despite having different topology have property values suggesting the presence of potential pharmacophore. It suggest the possibility to use this approach as a novel approach for computational drug design(AU)


Subject(s)
Humans , Medical Informatics Applications , Software Design , Pharmaceutical Preparations/standards , Molecular Dynamics Simulation/statistics & numerical data
19.
Rev. bras. anestesiol ; 65(3): 207-212, May-Jun/2015. tab, graf
Article in English | LILACS | ID: lil-748922

ABSTRACT

BACKGROUND AND OBJECTIVE: Postoperative pain treatment in mastectomy remains a major challenge despite the multimodal approach. The aim of this study was to investigate the analgesic effect of intravenous lidocaine in patients undergoing mastectomy, as well as the postoperative consumption of opioids. METHODS: After approval by the Human Research Ethics Committee of the Instituto de Medicina Integral Prof. Fernando Figueira in Recife, Pernambuco, a randomized, blind, controlled trial was conducted with intravenous lidocaine at a dose of 3 mg/kg infused over 1 h in 45 women undergoing mastectomy under general anesthesia. One patient from placebo group was. RESULTS: Groups were similar in age, body mass index, type of surgery, and postoperative need for opioids. Two of 22 patients in lidocaine group and three of 22 patients in placebo group requested opioid (p = 0.50). Pain on awakening was identified in 4/22 of lidocaine group and 5/22 of placebo group (p = 0.50); in the post-anesthetic recovery room in 14/22 and 12/22 (p = 0.37) of lidocaine and placebo groups, respectively. Pain evaluation 24 h after surgery showed that 2/22 and 3/22 patients (p = 0.50) of lidocaine and placebo groups, respectively, complained of pain. CONCLUSION: Intravenous lidocaine at a dose of 3 mg/kg administered over a period of an hour during mastectomy did not promote additional analgesia compared to placebo in the first 24 h, and has not decreased opioid consumption. However, a beneficial effect of intravenous lidocaine in selected and/or other therapeutic regimens patients cannot be ruled out. .


JUSTIFICATIVA E OBJETIVO: O tratamento da dor pós-operatória em mastectomia continua sendo um grande desafio apesar da abordagem multimodal. O objetivo deste estudo foi investigar o efeito analgésico da lidocaína intravenosa em pacientes submetidas a mastectomia, como também, o consumo de opioide pós-operatório. MÉTODOS: Após aprovação pelo comitê de ética e pesquisa em seres humanos do Instituto de Medicina Integral Prof. Fernando Figueira em Recife - Pernambuco foi realizado ensaio clínico aleatório encoberto placebo controlado com lidocaína intravenosa na dose de 3 mg/kg infundida em uma hora, em 45 mulheres submetidas a mastectomia sob anestesia geral. Excluída uma paciente do grupo placebo. RESULTADOS: Os grupos foram semelhantes quanto à idade, índice de massa corpórea, tipo de intervenção cirúrgica e necessidade de opioide no pós-operatório. Solicitaram opioide 2/22 pacientes nos grupos da lidocaína e 3/22 placebo (p = 0,50). Identificada a dor ao despertar em 4/22 no grupo lidocaína e 5/22 (p = 0,50) no grupo placebo; na sala de recuperação pós-anestésica em 14/22 e 12/22 (p = 0,37) nos grupos lidocaína e placebo respectivamente. Ao avaliar a dor 24 horas após o procedimento cirúrgico 3/22 e 2/22 (p = 0,50) das pacientes relataram dor em ambos os grupos respectivamente. CONCLUSÃO: A lidocaína intravenosa na dose de 3mg/kg administrada em um período de uma hora no transoperatório de mastectomia não promoveu analgesia adicional em relação ao grupo placebo nas primeiras 24 horas e não diminuiu o consumo de opioide. Contudo, um efeito benéfico da lidocaína intravenosa em pacientes selecionadas e/ou em outros regimes terapêuticos não pode ser descartado. .


JUSTIFICACIÓN Y OBJETIVO: El tratamiento del dolor postoperatorio en la mastectomía continúa siendo un gran reto a pesar del abordaje multimodal. El objetivo de este estudio fue investigar el efecto analgésico de la lidocaína intravenosa en pacientes sometidas a mastectomía, así como el consumo postoperatorio de opiáceos. MÉTODOS: Después de la aprobación por el Comité de Ética e Investigación en seres humanos del Instituto de Medicina Integral Prof. Fernando Figueira, en Recife, Pernambuco, se realizó un ensayo clínico aleatorizado, encubierto, placebo controlado con lidocaína intravenosa en una dosis de 3 mg/kg infundida en una hora, en 45 mujeres sometidas a mastectomía bajo anestesia general. Una paciente del grupo placebo fue excluida. RESULTADOS: Los grupos fueron similares en cuanto a la edad, índice de masa corporal, tipo de intervención quirúrgica y necesidad de opiáceos en el postoperatorio. Solicitaron opiáceos 2/22 pacientes en los grupos de la lidocaína y 3/22 placebo (p = 0,50). Fue identificado el dolor al despertar en 4/22 en el grupo lidocaína y 5/22 (p = 0,50) en el grupo placebo; en la sala de recuperación postanestésica en 14/22 y 12/22 (p = 0,37) en los grupos lidocaína y placebo, respectivamente. Al calcular el dolor 24 h después del procedimiento quirúrgico 3/22 y 2/22 (p = 0,50) de las pacientes relataron dolor en ambos grupos respectivamente. CONCLUSIÓN: La lidocaína intravenosa en una dosis de 3 mg/kg administrada en un período de una hora en el transoperatorio de mastectomía no generó analgesia adicional con relación al grupo placebo en las primeras 24 h y no disminuyó el consumo de opiáceos. Sin embargo, no puede ser descartado un efecto beneficioso de la lidocaína intravenosa en pacientes seleccionadas y/o en otros regímenes terapéuticos. .


Subject(s)
Humans , Metapneumovirus/genetics , Transcription, Genetic , Viral Proteins/chemistry , Amino Acid Sequence , Adenosine Monophosphate/metabolism , Crystallography, X-Ray , DNA , Edetic Acid/pharmacology , Molecular Dynamics Simulation , Molecular Sequence Data , Protein Binding , Protein Conformation , Protein Multimerization , Protein Stability , Protein Subunits/chemistry , RNA, Viral/metabolism , RNA, Viral/ultrastructure , Scattering, Small Angle , Solutions , Solvents , Viral Proteins/metabolism , Viral Proteins/ultrastructure , Zinc Fingers
20.
Indian J Biochem Biophys ; 2015 Feb; 52 (1): 23-28
Article in English | IMSEAR | ID: sea-157951

ABSTRACT

Crizotinib is the potential anticancer drug used for the treatment of non-small cell lung cancer (NSCLC) approved by FDA in 2011. The main target for the crizotinib is anaplastic lymphoma kinase (ALK). Evidences available indicate that double mutant ALK (L1196M and G1269A) confers resistance to crizotinib. However, how mutation confers drug resistance is not well-understood. Hence, in the present study, molecular dynamic (MD) simulation approach was employed to study the impact of crizotinib binding efficacy with ALK structures at a molecular level. Docking results indicated that ALK double mutant (L1196M and G1269A) significantly affected the binding affinity for crizotinib. Furthermore, MD studies revealed that mutant ALK-crizotinib complex showed higher deviation, higher fluctuation and decreased number of intermolecular H-bonds, when compared to the native ALK-crizotinib complex. These results may be immense importance for the molecular level understanding of the crizotinib resistance pattern and also for designing potential drug molecule for the treatment of lung cancer.


Subject(s)
Carcinoma, Non-Small-Cell Lung/drug therapy , Drug Resistance, Neoplasm/etiology , Molecular Dynamics Simulation/statistics & numerical data , Pyrazoles/pharmacology , Pyridines/pharmacology
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