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1.
J Comput Aided Mol Des ; 35(8): 871-882, 2021 08.
Artículo en Inglés | MEDLINE | ID: mdl-34181199

RESUMEN

Assessment of target druggability guided by search and characterization of hot spots is a pivotal step in early stages of drug-discovery. The raw output of FTMap provides the data to perform this task, but it relies on manual intervention to properly combine different sets of consensus sites, therefore allowing identification of hot spots and evaluation of strength, shape and distance among them. Thus, the user's previous experience on the target and the software has a direct impact on how data generated by FTMap server can be explored. DRUGpy plugin was developed to overcome this limitation. By automatically assembling and scoring all possible combinations of consensus sites, DRUGpy plugin provides FTMap users a straight-forward method to identify and characterize hot spots in protein targets. DRUGpy is available in all operating systems that support PyMOL software. DRUGpy promptly identifies and characterizes pockets that are predicted by FTMap to bind druglike molecules with high-affinity (druggable sites) or low-affinity (borderline sites) and reveals how protein conformational flexibility impacts on the target's druggability. The use of DRUGpy on the analysis of trypanothione reductases (TR), a validated drug target against trypanosomatids, showcases the usefulness of the plugin, and led to the identification of a druggable pocket in the conserved dimer interface present in this class of proteins, opening new perspectives to the design of selective inhibitors.


Asunto(s)
Descubrimiento de Drogas , Inhibidores Enzimáticos/farmacología , Modelos Moleculares , NADH NADPH Oxidorreductasas/antagonistas & inhibidores , Programas Informáticos , Sitios de Unión , Inhibidores Enzimáticos/química , Humanos , Ligandos , NADH NADPH Oxidorreductasas/química , NADH NADPH Oxidorreductasas/metabolismo , Unión Proteica , Conformación Proteica
2.
Nutr Metab Cardiovasc Dis ; 30(1): 141-150, 2020 01 03.
Artículo en Inglés | MEDLINE | ID: mdl-31757569

RESUMEN

BACKGROUND AND AIMS: Common beans (Phaseolus vulgaris L.) protein hydrolysate is a source of bioactive peptides with known health benefits. The aim of this study was to evaluate the effect of common bean protein hydrolysate on lipid metabolism and endothelial function in male adult BALB/c mice fed an atherogenic diet for nine weeks. METHODS AND RESULTS: Male adult mice were divided into three experimental groups (n = 12) and fed with normal control diet; atherogenic diet and atherogenic diet added with bean protein hydrolysate (700 mg/kg/day) for nine weeks. Food intake, weight gain, lipid profile, Atherogenic Index of Plasma, inflammation biomarkers and endothelial function were evaluated. APH group presented reduced feed intake, weight gain, lipid profile, tumor necrosis factor-α, angiotensin II (94% and 79%, respectively) and increased endothelial nitric oxide synthase (62%). CONCLUSIONS: Protein hydrolysate showed hypocholesterolemic activity preventing inflammation and dysfunction of vascular endothelium, in addition to decreasing oxidative stress, indicating an adjuvant effect on reducing atherogenic risk.


Asunto(s)
Anticolesterolemiantes/administración & dosificación , Aterosclerosis/prevención & control , Colesterol/sangre , Endotelio Vascular/efectos de los fármacos , Hipercolesterolemia/prevención & control , Phaseolus , Proteínas de Vegetales Comestibles/administración & dosificación , Hidrolisados de Proteína/administración & dosificación , Animales , Antiinflamatorios/administración & dosificación , Antioxidantes/administración & dosificación , Aterosclerosis/sangre , Aterosclerosis/etiología , Aterosclerosis/fisiopatología , Biomarcadores/sangre , Dieta Aterogénica , Modelos Animales de Enfermedad , Endotelio Vascular/metabolismo , Endotelio Vascular/fisiopatología , Hipercolesterolemia/sangre , Hipercolesterolemia/etiología , Hipercolesterolemia/fisiopatología , Mediadores de Inflamación/metabolismo , Masculino , Ratones Endogámicos BALB C , Óxido Nítrico Sintasa de Tipo III/metabolismo , Estrés Oxidativo/efectos de los fármacos
3.
Biochim Biophys Acta Gen Subj ; 1867(7): 130378, 2023 07.
Artículo en Inglés | MEDLINE | ID: mdl-37150227

RESUMEN

Tuberculosis (TB) is a leading cause of death worldwide. TB represents a serious public health threat, and it is characterized by high transmission rates, prevalence in impoverished regions, and high co-infection rates with HIV. Moreover, the serious side effects of long-term treatment that decrease patient adherence, and the emergence of multi-resistant strains of Mycobacterium tuberculosis, the causing agent of TBs, pose several challenges for its eradication. The search for a new TB treatment is necessary and urgent. Dihydroorotate dehydrogenase (DHODH) is responsible for the stereospecific oxidation of (S)-dihydroorotate (DHO) to orotate during the fourth and only redox step of the de novo pyrimidine nucleotide biosynthetic pathway. DHODH has been considered an attractive target against infectious diseases. As a first step towards exploiting DHODH as a drug target against TB, we performed a full kinetic characterization of both bacterial MtDHODH and its human ortholog (HsDHDOH) using both substrates coenzyme Q0 (Q0) and vitamin K3 (K3). MtDHODH follows a ping-pong mechanism of catalysis and shares similar catalytic parameters with the human enzyme. Serendipitously, Q0 was found to inhibit MtDHODH (KI (Q0) = 138 ± 31 µM). To the best of our knowledge, Q0 is the first non-orotate like dihydroorotate-competitive inhibitor for class 2 DHODHs ever described. Molecular dynamics simulations along with in silico solvent mapping allowed us to successfully probe protein flexibility and correlate it with the druggability of binding sites. Together, our results provide the starting point for the design of a new generation of potent and selective inhibitors against MtDHODH.


Asunto(s)
Mycobacterium tuberculosis , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH , Humanos , Dihidroorotato Deshidrogenasa , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/química , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/metabolismo , Mycobacterium tuberculosis/metabolismo , Sitios de Unión , Oxidación-Reducción
4.
Viruses ; 14(9)2022 Sep 05.
Artículo en Inglés | MEDLINE | ID: mdl-36146888

RESUMEN

The authors hereby request the inclusion of two authors (Olivia Teixeira and Maria Cristina Nonato) in the recently published article in Viruses entitled "Nucleocapsid (N) gene mutations of SARS-CoV-2 can affect real-time RT-PCR diagnostic and impact false-negative results" [...].

5.
Int J Biol Macromol ; 187: 964-975, 2021 Sep 30.
Artículo en Inglés | MEDLINE | ID: mdl-34310993

RESUMEN

Neospora caninum causes heavy losses related to abortions in bovine cattle. This parasite developed a complex defense redox system, composed of enzymes as glutathione reductase (GR). Methylene blue (MB) impairs the activity of recombinant form of Plasmodium GR and inhibits the parasite proliferation in vivo and in vitro. Likewise, MB and its derivatives inhibits Neospora caninum proliferation, however, whether the MB mechanism of action is correlated to GR function remains unclear. Therefore, here, N. caninum GR (NcGR) was characterized and its potential inhibitors were determined. NcGR was found in the tachyzoite cytosol and has a similar structure and sequence compared to its homologs. We verified the in vitro activity of rNcGR (875 nM) following NADPH absorbance at 340 nM (100 mM KH2PO4, pH 7.5, 1 mM EDTA, ionic strength: 600 mM, 25 °C). rNcGR exhibited a Michaelian behavior (Km(GSSG):0.10 ± 0.02 mM; kcat(GSSG):0.076 ± 0.003 s-1; Km(NADPH):0.006 ± 0.001 mM; kcat(NADPH): 0.080 ± 0.003 s-1). The IC50 of MB,1,9-dimethyl methylene blue, new methylene blue, and toluidine blue O on rNcGR activity were 2.1 ± 0.2 µM, 11 ± 2 µM, 0.7 ± 0.1 µM, and 0.9 ± 0.2 µM, respectively. Our results suggest the importance of NcGR in N. caninum biology and antioxidant mechanisms. Moreover, data presented here strongly suggest that NcGR is an important target of phenothiazinium dyes in N. caninum proliferation inhibition.


Asunto(s)
Coccidiostáticos/farmacología , Inhibidores Enzimáticos/farmacología , Glutatión Reductasa/efectos de los fármacos , Azul de Metileno/análogos & derivados , Neospora/efectos de los fármacos , Cloruro de Tolonio/farmacología , Animales , Citoplasma/enzimología , Glutatión Reductasa/genética , Glutatión Reductasa/metabolismo , Cinética , Masculino , Azul de Metileno/farmacología , Ratones Endogámicos BALB C , Neospora/enzimología , Neospora/genética , Neospora/crecimiento & desarrollo
6.
Viruses ; 13(12)2021 12 10.
Artículo en Inglés | MEDLINE | ID: mdl-34960743

RESUMEN

The current COVID-19 pandemic demands massive testing by Real-time RT-PCR (Reverse Transcription Polymerase Chain Reaction), which is considered the gold standard diagnostic test for the detection of the SARS-CoV-2 virus. However, the virus continues to evolve with mutations that lead to phenotypic alterations as higher transmissibility, pathogenicity or vaccine evasion. Another big issue are mutations in the annealing sites of primers and probes of RT-PCR diagnostic kits leading to false-negative results. Therefore, here we identify mutations in the N (Nucleocapsid) gene that affects the use of the GeneFinder COVID-19 Plus RealAmp Kit. We sequenced SARS-CoV-2 genomes from 17 positive samples with no N gene detection but with RDRP (RNA-dependent RNA polymerase) and E (Envelope) genes detection, and observed a set of three different mutations affecting the N detection: a deletion of 18 nucleotides (Del28877-28894), a substitution of GGG to AAC (28881-28883) and a frameshift mutation caused by deletion (Del28877-28878). The last one cause a deletion of six AAs (amino acids) located in the central intrinsic disorder region at protein level. We also found this mutation in 99 of the 14,346 sequenced samples by the Sao Paulo state Network for Pandemic Alert of Emerging SARS-CoV-2 variants, demonstrating the circulation of the mutation in Sao Paulo, Brazil. Continuous monitoring and characterization of mutations affecting the annealing sites of primers and probes by genomic surveillance programs are necessary to maintain the effectiveness of the diagnosis of COVID-19.


Asunto(s)
Prueba de Ácido Nucleico para COVID-19 , COVID-19/diagnóstico , Proteínas de la Nucleocápside de Coronavirus/genética , SARS-CoV-2/aislamiento & purificación , Brasil/epidemiología , COVID-19/epidemiología , ARN Polimerasa Dependiente de ARN de Coronavirus/genética , Cartilla de ADN , Reacciones Falso Negativas , Genoma Viral/genética , Humanos , Mutación , Fosfoproteínas/genética , ARN Viral/genética , SARS-CoV-2/genética
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