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Simple dialkyl pyrazole-3,5-dicarboxylates show in vitro and in vivo activity against disease-causing trypanosomatids.

2017

SUMMARYThe synthesis and antiprotozoal activity of some simple dialkyl pyrazole-3,5-dicarboxylates (compounds 2–6) and their sodium salts (pyrazolates) (compounds 7–9) against Trypanosoma cruzi, Leishmania infantum and Leishmania braziliensis are reported. In most cases the studied compounds showed, especially against the clinically significant amastigote forms, in vitro activities higher than those of the reference drugs (benznidazole for T. cruzi and glucantime for Leishmania spp.); furthermore, the low non-specific cytotoxicities against Vero cells and macrophages shown by these compounds led to good selectivity indexes, which are 8–72 times higher for T. cruzi amastigotes and 15–113 tim…

2019

Golgi α-mannosidase II (GMII) is a glycoside hydrolase playing a crucial role in the N-glycosylation pathway. In various tumour cell lines, the distribution of N-linked sugars on the cell surface is modified and correlates with the progression of tumour metastasis. GMII therefore is a possible molecular target for anticancer agents. Here, we describe the identification of a non-competitive GMII inhibitor using computer-aided drug design methods including identification of a possible allosteric binding site, pharmacophore search and virtual screening.

Et tu, Brute? Not Even Intracellular Mutualistic Symbionts Escape Horizontal Gene Transfer

2017

Many insect species maintain mutualistic relationships with endosymbiotic bacteria. In contrast to their free-living relatives, horizontal gene transfer (HGT) has traditionally been considered rare in long-term endosymbionts. Nevertheless, meta-omics exploration of certain symbiotic models has unveiled an increasing number of bacteria-bacteria and bacteria-host genetic transfers. The abundance and function of transferred loci suggest that HGT might play a major role in the evolution of the corresponding consortia, enhancing their adaptive value or buffering detrimental effects derived from the reductive evolution of endosymbionts' genomes. Here, we comprehensively review the HGT cases recor…

Food quality affects the expression of antimicrobial peptide genes upon simulated parasite attack in the larvae of greater wax moth

2017

Predator‐prey interactions are an important evolutionary force affecting the immunity of the prey. Parasitoids and mites pierce the cuticle of their prey, which respond by activating their immune system against predatory attacks. Immunity is a costly function for the organism, as it often competes with other life‐history traits for limited nutrients. We tested whether the expression of antimicrobial peptides (AMP) of the larvae of the greater wax moth Galleria mellonella (L.) (Lepidoptera: Pyralidae) changes as a consequence of insertion of a nylon monofilament, which acts like a synthetic parasite. The treatment was done for larvae grown on a high‐quality vs. a low‐quality diet. The expres…

Targeting of the Leishmania Mexicana cysteine protease CPB2.8 ΔCTE by decorated fused benzo[b] thiophene scaffold.

2016

A potent and highly selective anhydride-based inhibitor of Leishmania mexicana cysteine protease CPB2.8ΔCTE (IC50 = 3.7 μM) was identified. The details of the interaction of the ligand with the enzyme active site were investigated by NMR biomimetic experiments and docking studies. Results of inhibition assays, NMR and theoretical studies indicate that the ligand acts initially as a non-covalent inhibitor and later as an irreversible covalent inhibitor by chemoselective attack of CYS 25 thiolate to an anhydride carbonyl.

Entrapment and characterization of functional allosteric conformers of hemocyanin in sol–gel matrices

2016

Hemocyanins are giant oxygen transport proteins of molluscs and arthropods, which display high cooperativity and a complex pattern of conformations, generated by hierarchical allosteric interactions of their complex quaternary structure. A still unanswered question is the correlation between the functional properties of the postulated conformers and structural features that govern their oxygen binding, such as metal complex coordination. In this study we focus on the dodecameric hemocyanin of the crustacean Carcinus aestuarii, with the aim to obtain a functional and structural characterization of the individual conformational states giving rise to cooperativity, by entrapping hemocyanin int…

Revealing the Origin of the Efficiency of the De Novo Designed Kemp Eliminase HG-3.17 by Comparison with the Former Developed HG-3

2017

The design of new biocatalysts is a goal in biotechnology to improve the rate, selectivity and environmental impact of industrial chemical processes. In this regard, the use of computational techniques has provided valuable assistance in the design of new enzymes with remarkable catalytic activity. In this paper, hybrid QM/MM molecular dynamics simulations have allowed insights to be gained on the origin of the limited efficiency of a computationally designed enzyme for the Kemp elimination; the HG-3. Comparison of results derived from this enzyme with those of a more evolved protein containing additional point mutations, HG-3.17, rendered important information that should be taken into acc…

2018

The origin of substrate preference in promiscuous enzymes was investigated by enzyme isotope labelling of the alcohol dehydrogenase from Geobacillus stearothermophilus (BsADH). At physiological temperature, protein dynamic coupling to the reaction coordinate was insignificant. However, the extent of dynamic coupling was highly substrate-dependent at lower temperatures. For benzyl alcohol, an enzyme isotope effect larger than unity was observed, whereas the enzyme isotope effect was close to unity for isopropanol. Frequency motion analysis on the transition states revealed that residues surrounding the active site undergo substantial displacement during catalysis for sterically bulky alcohol…

Isolation and Analysis of Fasciola hepatica Extracellular Vesicles

2020

The finding of extracellular vesicles (EVs) as important players in parasite-parasite and host-parasite communications has led to an increasing number of reports in the literature. Different protocols have been developed for isolation and further characterization of EVs from parasitic helminths. In this chapter, we describe step by step procedures to isolate EVs secreted by Fasciola hepatica adults in culture, which could be also applied for other developmental stages of the parasite, as well as EVs present in plasma and urine. Along with classical isolation methods like differential ultracentrifugation, and more recent techniques like size exclusion chromatography (SEC), here we also refer…

Description of embryonic development and ultrastructure in miracidia ofCardiocephaloides longicollis(Digenea, Strigeidae) in relation to active host …

2017

The functional ultrastructure and embryonic development of miracidia in naturally released eggs of the trematode Cardiocephaloides longicollis were studied using light and transmission electron microscopy. This species has operculated eggs and embryogenesis occurs in the marine environment before an actively infecting ciliated miracidium hatches. Six different developmental stages were identified. The lack of pores in the eggshell indicates its impermeability and the miracidium's dependency on glycogen nutritive reserves, contained in numerous vitellocytes in early embryos. As the development advances, these merge into larger vitelline vacuoles that encircle the miracidium and may aid its h…