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The air-breathing magur catfish (Clarias magur) are frequently challenged with high environmental pollutants, including that of various metal nanoparticles (NPs) in their natural habitats. Heat shock proteins (HSPs) are essential molecular chaperones for preserving intracellular protein homeostasis in eukaryotic cells. In aquatic animals, HSPs are known to play important defensive roles associated with various environmental stress-related cellular damages. In the present investigation, we characterized the molecular and structural organization of distinct HSPs and their potential induction of HSP genes in multiple magur catfish tissues while exposed to ZnO NPs for 14 days. The sequence alignment of four HSP genes (hsp70, hsc70, hsp90a, and hsp90b) of magur catfish demonstrated evolutionary parallels with bony fishes and total conservation of active sites across the amphibia, fish, and mammals. From the architectural analysis of HSP70, HSC70, HSP90a, and HSP90b proteins, a structural similarity with mammals was observed, suggesting the functional resemblances of the studied HSPs in chaperone mechanisms. In the examined tissues, the mRNAs of HSP genes expressed constitutively. Exposure of C. magur to ZnO NPs (10 mg/L) in situ led to a considerable increase in the levels of mRNAs for several HSP genes and translated proteins, with HSP70 exhibiting the highest level of expression. Thus, it can be contemplated that HSPs may be involved in defending the magur catfish against the ZnO NP- and other metal NP-mediated cellular damages. The results provide new insights into the involvement of HSP machinery during adaptation to the ZnO NP-induced stress in magur catfish.
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Continuous attempts have been made to pinpoint candidate vaccine molecules and evaluate their effectiveness in order to commercialise such vaccines for the treatment of tropical fascioliasis in livestock. The pathophysiology of fascioliasis can be related to liver damage brought on by immature flukes that migrate and feed, as well as immunological reactions to chemicals produced by the parasites and alarm signals brought on by tissue damage. Future research should, in our opinion, concentrate on the biology of invasive parasites and the resulting immune responses, particularly in the early stages of infection. The goal of the current study was to use the calcium-binding proteins from F. gigantica to create a multi-epitope subunit vaccine. The adjuvant, B-cell epitopes, CTL epitopes, and HTL epitopes that make up the vaccine construct are all connected by certain linkers. The antigenicity, allergenicity, and physiochemical properties of the vaccine construct were examined. The vaccine construct was docked with toll-like receptor 2, and simulations of the molecular dynamics of the complex's stability, interaction, and dynamics were run. After performing in silico cloning and immunosimulation, it was discovered that the construct was suitable for further investigation. New vaccination technologies and adjuvant development are advancing our food safety procedures since vaccines are seen as safe and are accepted by the user community. This research is also applicable to the F. hepatica system.
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Fasciola , Fascioliasis , Animales , Fascioliasis/prevención & control , Calcio , Vacunas de Subunidad/química , Epítopos de Linfocito B , Epítopos de Linfocito T , Biología Computacional/métodos , Simulación del Acoplamiento MolecularRESUMEN
Thioredoxin glutathione reductase (TGR), a potential anthelminthic drug target causes NADPH-dependent transfer of electrons to both thioredoxins and glutathione systems. In the present study, we showed that a single point mutation conserved at Arg451 position is critical for maintaining the structure-function of FgTGR. The current biochemical results showed that R451A mutation significantly decreases both oxidoreductase activities (glutathione reductase and thioredoxin reductase) of the enzyme. Computational analyses using molecular dynamics simulation provided an in-depth insight into the structural alterations caused as a result of the mutation. Furthermore, the different regions of the mutant FgTGR structure were found to be altered in flexibility/rigidity as a result of the mutation. This led to mutant-specific conformational alterations and dominant differential motions that contributed to the abrogated function of mutant FgTGR. These results were confirmed using GdnHCl-induced denaturation-based stability studies. Moreover, mutation reduced the free energy of stabilization of the protein, thereby destabilizing the mutant protein structure. Therefore, these findings displayed differential dynamics in the FgTGR structure and highlighted the relevance of residue-level interactions in the protein. Thus, the current study provided a basis for exploiting regions other than the active site of TGR for inhibitory effect and development of novel antihelminthics.
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Arginina/química , Proteínas del Helminto/química , Complejos Multienzimáticos/química , NADH NADPH Oxidorreductasas/química , Animales , Dominio Catalítico , Fasciola/enzimología , Proteínas del Helminto/genética , Proteínas del Helminto/metabolismo , Simulación de Dinámica Molecular , Complejos Multienzimáticos/genética , Complejos Multienzimáticos/metabolismo , NADH NADPH Oxidorreductasas/genética , NADH NADPH Oxidorreductasas/metabolismo , Mutación Puntual , Análisis de Componente Principal , Estabilidad Proteica , Estructura Secundaria de Proteína/genética , Desplegamiento Proteico , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , TermodinámicaRESUMEN
The inherent ability of the cancer cells to resist chemotherapeutic agents is a major challenge in drug discovery. Chemotherapy is one of the most widely used treatment methods for cancer, but due to multidrug resistance (MDR) development in cancer cells, the healing procedure often fails. Various factors impart cancer resistance to cells; among them, P-glycoprotein (P-gp) overexpression is directly linked to MDR in cancer cells. P-gp leads to the efflux of drug molecules to the extracellular space. Several molecules have been reported to inhibit the P-gp activity. Among them, quercetin has revealed a great potential to modulate P-gp activity. However, the mechanistic understanding of quercetin induced modulation is not entirely elucidated. In the present work, we showed that quercetin binds in the interacting region between the transmembrane domain and nucleotide-binding domain out of the three plausible binding sites of P-gp and restrict the conformational change from inward- to outward-facing conformation of P-gp. Due to the absence of the inward-facing structure of human P-gp, we first modeled an inward-facing P-gp structure. Using molecular docking, the interacting residues of P-gp were identified, and the stability and interaction dynamics of the complex were studied using molecular dynamics simulation. Our work reveals the mechanistic understanding of quercetin induced modulation of P-gp and indicates its importance in cancer treatment.Communicated by Ramaswamy H. Sarma.
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Neoplasias , Quercetina , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/química , Línea Celular Tumoral , Resistencia a Múltiples Medicamentos , Resistencia a Antineoplásicos , Humanos , Simulación del Acoplamiento Molecular , Nucleótidos/metabolismo , Quercetina/farmacología , Transducción de SeñalRESUMEN
The tropical liver fluke, Fasciola gigantica is a food-borne parasite responsible for the hepatobiliary disease fascioliasis. The recent completion of F. gigantica genome sequencing by our group has provided a platform for the systematic analysis of the parasite genome. Eukaryotic protein kinases (ePKs) are regulators of cellular phosphorylation. In the present study, we used various computational and bioinformatics tools to extensively analyse the ePKs in F. gigantica (FgePKs) genome. A total of 455 ePKs were identified that represent ~2% of the parasite genome. Out of these, 214 ePKs are typical kinases (Ser/Thr- and Tyr-specific ePKs), and 241 were other kinases. Several FgePKs were found to possess unusual domain architectures, which suggests the diverse nature of the proteins that can be exploited for designing novel inhibitors. 115 kinases showed <35% query coverage when compared to human ePKs highlighting significant divergences in their respective kinomes, further providing a platform for novel structure-based drug designing. This study provides a platform that may open new avenues into our understanding of helminth biochemistry and drug discovery.
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Células Eucariotas/enzimología , Fasciola/genética , Genoma de los Helmintos/genética , Estudio de Asociación del Genoma Completo/métodos , Proteínas del Helminto/genética , Proteínas Quinasas/genética , Animales , Biología Computacional/métodos , Fasciola/enzimología , Fasciola/fisiología , Fascioliasis/parasitología , Proteínas del Helminto/clasificación , Proteínas del Helminto/metabolismo , Humanos , Isoenzimas/clasificación , Isoenzimas/genética , Isoenzimas/metabolismo , Familia de Multigenes/genética , Fosforilación , Filogenia , Proteínas Quinasas/clasificación , Proteínas Quinasas/metabolismoRESUMEN
Visceral Leishmaniasis (VL) is an insect-borne neglected disease caused by the protozoan parasite Leishmania donovani. In the absence of a commercial vaccine against VL, chemotherapy is currently the only option used for the treatment of VL. Vaccination has been considered as the most effective and powerful tool for complete eradication and control of infectious diseases. In this study, we aimed to design a peptide-based vaccine against L. donovani using immuno-bioinformatic tools. We identified 6 HTL, 18 CTL, and 25 B-cell epitopes from three hypothetical membrane proteins of L. donovani. All these epitopes were used to make a vaccine construct along with linkers. An adjuvant was also added at the N-terminal to enhance its immunogenicity. After that, we checked the quality of this vaccine construct and found that it is nontoxic, nonallergic, and thermally stable. A 3D structure of the vaccine construct was also generated by homology modeling to evaluate its interaction with innate immune receptors (TLR). Molecular docking was performed, which confirmed its binding with a toll-like receptor-2 (TLR-2). The stability of vaccine-TLR-2 complex and underlying interactions were evaluated using molecular dynamic simulation. Lastly, we carried out in silico cloning to check the expression of the final designed vaccine. The designed vaccine construct needs further experimental and clinical investigations to develop it as a safe and effective vaccine against VL infection.
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Epítopos de Linfocito B/inmunología , Epítopos de Linfocito T/inmunología , Leishmania , Vacunas Antiprotozoos/inmunología , Leishmaniasis/prevención & control , Simulación del Acoplamiento Molecular , Vacunas de Subunidad/inmunologíaRESUMEN
Fascioliasis, a neglected foodborne disease caused by liver flukes (genus Fasciola), affects more than 200 million people worldwide. Despite technological advances, little is known about the molecular biology and biochemistry of these flukes. We present the draft genome of Fasciola gigantica for the first time. The assembled draft genome has a size of â¼1.04 Gb with an N50 and N90 of 129 and 149 kb, respectively. A total of 20â¯858 genes were predicted. The de novo repeats identified in the draft genome were 46.85%. The pathway included all of the genes of glycolysis, Krebs cycle, and fatty acid metabolism but lacked the key genes of the fatty acid biosynthesis pathway. This indicates that the fatty acid required for survival of the fluke may be acquired from the host bile. It may be hypothesized that the relatively larger F. gigantica genome did not evolve through genome duplications but rather is interspersed with many repetitive elements. The genomic information will provide a comprehensive resource to facilitate the development of novel interventions for fascioliasis control.
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Granular cell tumors are rare soft tissue neoplasms, among which only 2% are malignant, arising from nervous tissue. Here we present a case of a large esophageal granular cell tumor with benign histopathological features which metastasized to the liver, but showing on positron emission tomography-computerized tomography standardized uptake value suggestive of a benign lesion.