Papers by Maikel Izquierdo Rivero
PLOSNTD, 2024
Background Chagas disease is caused by the protozoan parasite Trypanosoma cruzi and leads tõ 10,0... more Background Chagas disease is caused by the protozoan parasite Trypanosoma cruzi and leads tõ 10,000 deaths each year. Nifurtimox and benznidazole are the only two drugs available but have significant adverse effects and limited efficacy. New chemotherapeutic agents are urgently required. Here we identified inhibitors of the acidic M17 leucyl-aminopeptidase from T. cruzi (LAPTc) that show promise as novel starting points for Chagas disease drug discovery.
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ECIMED, 2010
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Citrifrut, 2013
El mango es una especie que ocupa en Cuba el segundo lugar en la producción de frutas, después de... more El mango es una especie que ocupa en Cuba el segundo lugar en la producción de frutas, después de los cítricos, por su buena adaptación a las condiciones edafoclimáticas del país y la aceptación de sus frutos para el consumo, tanto en fresco, como elaborados en forma de jugos, mermeladas, tajadas en almíbar y compotas. La dinámica del mercado de mango en la actualidad, está exigiendo nuevas estrategias en los programas de mejoramiento fundamentalmente dirigidas a la obtención de nuevos genotipos que respondan a las demandas comerciales. La estrecha base genética encontrada en el cultivo, a partir de la caracterización molecular realizada en 30 de los principales cultivares, indicó la necesidad de implementar otros métodos de mejoramiento diferentes a la selección directa, que dieran lugar a nuevos genotipos. El presente trabajo tuvo como objetivo la caracterización de una progenie de mango obtenida por cruzamiento abierto. El progenitor femenino seleccionado fue el cv. 'Julie' caracterizado por su bajo porte. Los resultados mostraron que la mayor diversidad se encontró en las variables relacionadas con el fruto, No hubo diferencias significativas con respecto al porte de las plantas, solo tres de las selecciones obtenidas presentaron alturas de la planta menores al resto de la progenie. Una de ellas se distinguió por presentar además semillas poliembriónicas, carácter ausente en el cv. 'Julie', causa principal que ha limitado su utilización como portainjerto. palabras clave: mango, Mangifera indica L., diversidad morphoagronomic diversity of a mango (Mangifera indica L.) progeny cv. 'Julie' obtained by open crossing aBsTracT Mango is a species that occupies the second place in Cuba in fruits production after citrus fruits, for its good adaptation to the edaphoclimatic conditions of the country and by the acceptance of its fruits for the consumption in fresh, as well as elaborated in juices, marmalades, slices in syrup and preserves. The dynamics of mango market at the present time is demanding new strategies in the programs of improvement mainly directed to obtaining new genotypes that respond to the commercial demands. The narrow genetic base found in the culture, starting from the molecular characterization carried out in 30 of the main cultivars, indicated the necessity to implement other methods of improvement different to the direct selection that gave place to new genotypes. The present work had as objective the characterization of a mango progeny obtained by open crossing. The selected female progenitor was the cv. 'Julie' characterized by their low erecting. The results showed that the biggest diversity was in the variables related with the fruit, there were not significant differences with regard to the plants behavior, only three of the obtained selections presented plant heights minor from the rest of the progeny. One of them was distinguished by presenting polyembryonic seed; absent character in the cv. 'Julie' which is the main cause that has limited its use as rootstock.
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RCCB, 2017
Caracterización cualitativa de una progenie de mango (Mangifera indica L.) obtenida por polinizac... more Caracterización cualitativa de una progenie de mango (Mangifera indica L.) obtenida por polinización abierta Qualitative characterization of a mango progeny (Mangifera indica L.) obtained by open pollination
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RCCB, 2018
Molecular markers have proven to be powerful tools in research related to diversity, variability ... more Molecular markers have proven to be powerful tools in research related to diversity, variability and improvement of economically important tropical crops. Retrotransposons are ubiquitous and major components of eukaryotic genomes reaching high copy numbers in most plant species. The main goal of this study was to analyze the genetic relationship among 34 mango (Mangifera indica L) accessions using three previously selected Inverse Sequence-Tagged Repeats (ISTR) markers shown to be the the most informative ISTR combination for this crop. These ISTR combinations generated 40 polymorphic fragments, demonstrating its usefulness for the estimation of the genetic diversity in mango accessions. The primers combination F2 / B2B and F5 / B2B showed 100% of polymorphism bands, while 93.75% of polymorphism was detected with the primers F3 / B3. On the other hand, Genotype-6 (Gen-6) did not show genetic similarity with any of the other studied genotypes. This study provides further support for these ISTR combinations as a potential tool to identify promissory genotypes in this crop´s progenies. This result also suggests the possibility to use genotype 6 as a mother plant in the second stage of mango progeny improvement.
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European Journal of Medicinal Chemistry, 2019
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The Protein Journal, 2019
The M17 leucyl-aminopeptidase of Trypanosoma cruzi (LAPTc) is a novel drug target for Chagas dise... more The M17 leucyl-aminopeptidase of Trypanosoma cruzi (LAPTc) is a novel drug target for Chagas disease. The objective of this work was to obtain recombinant LAPTc (rLAPTc) in Escherichia coli. A LAPTc gene was designed, optimized for its expression in E. coli, synthesized and cloned into the vector pET-19b. Production of rLAPTc in E. coli BL21(DE3) pLysS, induced for 20 h at 25 °C with 1 mM IPTG, yielded soluble rLAPTC that was catalytically active. The rLAPTc enzyme was purified in a single step by IMAC. The recombinant protein was obtained with a purity of 90% and a volumetric yield of 90 mg per liter of culture. The enzymatic activity has an optimal pH of 9.0, and preference for Leu-p-nitroanilide (appK M = 74 µM, appk cat = 4.4 s −1). The optimal temperature is 50 °C, and the cations Mg 2+ , Cd 2+ , Ba 2+ , Ca 2+ and Zn 2+ at 4 mM inhibited the activity by 60% or more, while Mn 2+ inhibited by only 15% and addition of Co 2+ activated by 40%. The recombinant enzyme is insensitive toward the protease inhibitors PMSF, TLCK, E-64 and pepstatin A, but is inhibited by EDTA and bestatin. Bestatin is a non-competitive inhibitor of the enzyme with a K i value of 881 nM. The enzyme is a good target for inhibitor identification.
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RCCB, 2019
Structure-activity relationship of the inhibition of M1aminopeptidases from Escherichia coli (ePe... more Structure-activity relationship of the inhibition of M1aminopeptidases from Escherichia coli (ePepN) and Plasmodium falciparum (PfA-M1) by bestatin-derived peptidomimetics Relación estructura-actividad de la inhibición de aminopeptidasas M1 de Escherichia coli (ePepN) y Plasmodium falciparum (PfA-M1) por peptidomiméticos derivados de la bestatina
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SLAS Discovery, 2020
Leucyl aminopeptidases (LAPs) are involved in multiple cellular functions, which, in the case of ... more Leucyl aminopeptidases (LAPs) are involved in multiple cellular functions, which, in the case of infectious diseases, includes participation in the pathogen-host cell interface and pathogenesis. Thus, LAPs are considered good candidate drug targets, and the major M17-LAP from Trypanosoma cruzi (LAPTc) in particular is a promising target for Chagas disease. To exploit LAPTc as a potential target, it is essential to develop potent and selective inhibitors. To achieve this, we report a highthroughput screening method for LAPTc. Two methods were developed and optimized: a Leu-7-amido-4-methylcoumarinbased fluorogenic assay and a RapidFire mass spectrometry (RapidFire MS)-based assay using the LSTVIVR peptide as substrate. Compared with a fluorescence assay, the major advantages of the RapidFire MS assay are a greater signal-to-noise ratio as well as decreased consumption of enzyme. RapidFire MS was validated with the broad-spectrum LAP inhibitors bestatin (IC 50 ¼ 0.35 mM) and arphamenine A (IC 50 ¼ 15.75 mM). We suggest that RapidFire MS is highly suitable for screening for specific LAPTc inhibitors.
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Microbiall Cell, 2021
Several microbial metalo-aminopeptidases are emerging as novel targets for the treatment of human... more Several microbial metalo-aminopeptidases are emerging as novel targets for the treatment of human infectious diseases. Some of them are well validated as targets and some are not; some are essential enzymes and others are important for virulence and pathogenesis. For another group, it is not clear if their enzymatic activity is involved in the critical functions that they mediate. But one aspect has been established: they display relevant roles in bacteria and protozoa that could be targeted for therapeutic purposes. This work aims to describe these biological functions for several microbial metalo-aminopeptidases.
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Life, 2021
Chagas disease, caused by the kinetoplastid parasite Trypanosoma cruzi, is a human tropical illne... more Chagas disease, caused by the kinetoplastid parasite Trypanosoma cruzi, is a human tropical illness mainly present in Latin America. The therapies available against this disease are far from ideal. Proteases from pathogenic protozoan have been considered as good drug target candidates. T. cruzi acidic M17 leucyl-aminopeptidase (TcLAP) mediates the major parasite's leucyl-aminopeptidase activity and is expressed in all parasite stages. Here, we report the inhibition of TcLAP (IC50 = 66.0 ± 13.5 μM) by the bestatin-like peptidomimetic KBE009. This molecule also inhibited the proliferation of T. cruzi epimastigotes in vitro (EC50 = 28.1 ± 1.9 μM) and showed selectivity for the parasite over human dermal fibroblasts (selectivity index: 4.9). Further insight into the specific effect of KBE009 on T. cruzi was provided by docking simulation using the crystal structure of TcLAP and a modeled human orthologous, hLAP3. The TcLAP-KBE009 complex is more stable than its hLAP3 counterpart. KBE009 adopted a better geometrical shape to fit into the active site of TcLAP than that of hLAP3. The drug-likeness and lead-likeness in silico parameters of KBE009 are satisfactory. Altogether, our results provide an initial insight into KBE009 as a promising starting point compound for the rational design of drugs through further optimization.
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Protein Expression and Purification, 2021
The Leishmania major leucyl-aminopeptidase (LAPLm), a member of the M17 family of proteases, is a... more The Leishmania major leucyl-aminopeptidase (LAPLm), a member of the M17 family of proteases, is a potential drug target for treatment of leishmaniasis. To better characterize enzyme properties, recombinant LAPLm (rLAPLm) was expressed in Escherichia coli. A LAPLm gene was designed, codon-optimized for expression in E. coli, synthesized and cloned into the pET-15b vector. Production of rLAPLm in E. coli Lemo21(DE3), induced for 4 h at 37 • C with 400 μM IPTG and 250 μM L-rhamnose, yielded insoluble enzyme with a low proportion of soluble and active protein, only detected by an anti-His antibody-based western-blot. rLAPLm was purified in a single step by immobilized metal ion affinity chromatography. rLAPLm was obtained with a purity of ~10% and a volumetric yield of 2.5 mg per liter, sufficient for further characterization. The aminopeptidase exhibits optimal activity at pH 7.0 and a substrate preference for Leu-p-nitroanilide (appK M = 30 μM, appk cat = 14.7 s − 1). Optimal temperature is 50 • C, and the enzyme is insensitive to 4 mM Co 2+ , Mg 2+ , Ca 2+ and Ba 2+. However, rLAPLm was activated by Zn 2+ , Mn 2+ and Cd 2+ but is insensitive towards the protease inhibitors PMSF, TLCK, E− 64 and pepstatin A, being inhibited by EDTA and bestatin. Bestatin is a potent, non-competitive inhibitor of the enzyme with a K i value of 994 nM. We suggest that rLAPLm is a suitable target for inhibitor identification.
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Background: Human infectious diseases caused by bacteria are a worldwide health problem due to th... more Background: Human infectious diseases caused by bacteria are a worldwide health problem due to the increased resistance of these microorganisms to conventional antibiotics. For this reason, the identification of novel molecular targets and the discovery of new antibacterial compounds are urgently required. Metalo-aminopeptidases are promising targets in bacterial infections. They participate in crucial processes for bacterial growth and pathogenesis, such as protein and peptide degradation to supply amino acids, protein processing, access to host tissues, cysteine supply for redox control, transcriptional regulation, site-specific DNA recombination, and hydrogen sulfide production. Although several of these enzymes are not essential, they are required for virulence and maximal growth in conditions of nutrient limitation and high temperatures. Objective: In this review, we describe the structural, functional, and kinetic properties of some examples of bacterial metalo-aminopeptidases, in the context of their use as antibacterial targets. In addition, we present some inhibitors reported for these enzymes. Conclusion: It is necessary to conduct a meticulous work to validate these peptidases as good/bad targets and to identify inhibitors with potential therapeutic use.
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Background: Parasitic human infectious diseases are a worldwide health problem due to the increas... more Background: Parasitic human infectious diseases are a worldwide health problem due to the increased resistance to conventional drugs. For this reason, the identification of novel molecular targets and the discovery of new chemotherapeutic agents are urgently required. Metalo-aminopeptidases are promising targets in parasitic infections. They participate in crucial processes for parasite growth and pathogenesis. Objective: In this review, we describe the structural, functional and kinetic properties, and inhibitors, of several parasite metalo-aminopeptidases, for their use as targets in parasitic diseases. Conclusion: Plasmodium falciparum M1 and M17 aminopeptidases are essential enzymes for parasite development, and M18 aminopeptidase could be involved in hemoglobin digestion and erythrocyte invasion and egression. Trypanosoma cruzi, T. brucei and Leishmania major acidic M17 aminopeptidases can play a nutritional role. T. brucei basic M17 aminopeptidase down-regulation delays the cytokinesis. The inhibition of Leishmania basic M17 aminopeptidase could affect parasite viability. L. donovani methionyl aminopeptidase inhibition prevents apoptosis but not the parasite death. Decrease in Acanthamoeba castellanii M17 aminopeptidase activity produces cell wall structural modifications and encystation inhibition. Inhibition of Babesia bovis growth is probably related to the inhibition of the parasite M17 aminopeptidase, probably involved in host hemoglobin degradation. Schistosoma mansoni M17 aminopeptidases inhibition may affect parasite development, since they could participate in hemoglobin degradation, surface membrane remodeling and eggs hatching. Toxoplasma gondii M17 aminopeptidase inhibition could attenuate parasite virulence, since it is apparently involved in the hydrolysis of cathepsin Cs-or proteasome-produced dipeptides and/or cell attachment/invasion processes. These data are relevant to validate these enzymes as targets.
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PLOS NEGLECTED TROPICAL DISEASES, 2024
Background Chagas disease is caused by the protozoan parasite Trypanosoma cruzi and leads tõ 10,0... more Background Chagas disease is caused by the protozoan parasite Trypanosoma cruzi and leads tõ 10,000 deaths each year. Nifurtimox and benznidazole are the only two drugs available but have significant adverse effects and limited efficacy. New chemotherapeutic agents are urgently required. Here we identified inhibitors of the acidic M17 leucyl-aminopeptidase from T. cruzi (LAPTc) that show promise as novel starting points for Chagas disease drug discovery.
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Papers by Maikel Izquierdo Rivero