BackgroundThe yellow fever mosquito, Aedes aegypti, is the primary vector for the viruses that cause yellow fever, mostly in tropical regions of Africa and in parts of South America, and human dengue, which infects 100 million people yearly in the tropics and subtropics. A better understanding of the structural biology of olfactory proteins may pave the way for the development of environmentally-friendly mosquito attractants and repellents, which may ultimately contribute to reduction of mosquito biting and disease transmission.MethodologyPreviously, we isolated and cloned a major, female-enriched odorant-binding protein (OBP) from the yellow fever mosquito, AaegOBP1, which was later inadvertently renamed AaegOBP39. We prepared recombinant samples of AaegOBP1 by using an expression system that allows proper formation of disulfide bridges and generates functional OBPs, which are indistinguishable from native OBPs. We crystallized AaegOBP1 and determined its three-dimensional structure at 1.85 Å resolution by molecular replacement based on the structure of the malaria mosquito OBP, AgamOBP1, the only mosquito OBP structure known to date.ConclusionThe structure of AaegOBP1 ( = AaegOBP39) shares the common fold of insect OBPs with six α-helices knitted by three disulfide bonds. A long molecule of polyethylene glycol (PEG) was built into the electron-density maps identified in a long tunnel formed by a crystallographic dimer of AaegOBP1. Circular dichroism analysis indicated that delipidated AaegOBP1 undergoes a pH-dependent conformational change, which may lead to release of odorant at low pH (as in the environment in the vicinity of odorant receptors). A C-terminal loop covers the binding cavity and this “lid” may be opened by disruption of an array of acid-labile hydrogen bonds thus explaining reduced or no binding affinity at low pH.

The development of cruzain inhibitors has been driven by the urgent need to develop novel and more effective drugs for the treatment of Chagas' disease. Herein, we report the lead optimization of a class of noncovalent cruzain inhibitors, starting from an inhibitor previously cocrystallized with the enzyme (K i = 0.8 μM). With the goal of achieving a better understanding of the structure−activity relationships, we have synthesized and evaluated a series of over 40 analogues, leading to the development of a very promising competitive inhibitor (8r, IC 50 = 200 nM, K i = 82 nM). Investigation of the in vitro trypanocidal activity and preliminary cytotoxicity revealed the potential of the most potent cruzain inhibitors in guiding further medicinal chemistry efforts to develop drug candidates for Chagas' disease.

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Endophytic actinobacteria from the Brazilian medicinal plant Lychnophora ericoides were isolated for the first time, and the biological potential of their secondary metabolites was evaluated. A phylogenic analysis of isolated actinobacteria was accomplished with 16S rRNA gene sequencing, and the predominance of the genus Streptomyces was observed. All strains were cultured on solid rice medium, and ethanol extracts were evaluated with antimicrobial and cytotoxic assays against cancer cell lines. As a result, 92% of the extracts showed a high or moderate activity against at least one pathogenic microbial strain or cancer cell line. Based on the biological and chemical analyses of crude extracts, three endophytic strains were selected for further investigation of their chemical profiles. Sixteen compounds were isolated, and 3-hydroxy-4-methoxybenzamide (9) and 2,3-dihydro-2,2-dimethyl-4(1H)-quinazolinone (15) are reported as natural products for the first time in this study. The biological activity of the pure compounds was also assessed. Compound 15 displayed potent cytotoxic activity against all four tested cancer cell lines. Nocardamine (2) was only moderately active against two cancer cell lines but showed strong activity against Trypanosoma cruzi. Our results show that endophytic actinobacteria from L. ericoides are a promising source of bioactive compounds.

BackgroundViolacein is a deep violet compound that is produced by a number of bacterial species. It is synthesized from tryptophan by a pathway that involves the sequential action of 5 different enzymes (encoded by genes vioA to vioE). Violacein has antibacterial, antiparasitic, and antiviral activities, and also has the potential of inducing apoptosis in certain cancer cells.ResultsHere, we describe the construction of a series of plasmids harboring the complete or partial violacein biosynthesis operon and their use to enable production of violacein and deoxyviolacein in E.coli. We performed in vitro assays to determine the biological activity of these compounds against Plasmodium, Trypanosoma, and mammalian cells. We found that, while deoxyviolacein has a lower activity against parasites than violacein, its toxicity to mammalian cells is insignificant compared to that of violacein.ConclusionsWe constructed E. coli strains capable of producing biologically active violacein and related compounds, and propose that deoxyviolacein might be a useful starting compound for the development of antiparasite drugs.Electronic supplementary materialThe online version of this article (10.1186/s12896-018-0428-z) contains supplementary material, which is available to authorized users.

Chagas disease causes ~10,000 deaths each year, mainly in Latin America, where it is endemic. The currently available chemotherapeutic agents are ineffective in the chronic stage of the disease, and the lack of pharmaceutical innovation for Chagas disease highlights the urgent need for the development of new drugs. The enzyme cruzain, the main cysteine protease of Trypanosoma cruzi, has been explored as a validated molecular target for drug discovery. Herein, the design, molecular modeling studies, synthesis, and biological evaluation of cyclic imides as cruzain inhibitors are described. Starting with a micromolar-range cruzain inhibitor (3a, IC50 = 2.2 μM), this molecular optimization strategy resulted in the nanomolar-range inhibitor 10j (IC50 = 0.6 μM), which is highly active against T. cruzi intracellular amastigotes (IC50 = 1.0 μM). Moreover, most compounds were selective toward T. cruzi over human fibroblasts, which were used as host cells, and are less toxic to hepatic cells than the marketed drug benznidazole. This study enabled the discovery of novel chemical diversity and established robust structure-activity relationships to guide the design of optimized cruzain inhibitors as new trypanocidal agents.

A virtual screening conducted with nearly 4 000 000 compounds from lead-like and fragment-like subsets enabled the identification of a small-molecule inhibitor (1) of the Trypanosoma cruzi cruzain enzyme, a validated drug target for Chagas disease. Subsequent comprehensive structure-based drug design and structure−activity relationship studies led to the discovery of carbamoyl imidazoles as potent, reversible, and competitive cruzain inhibitors. The most potent carbamoyl imidazole inhibitor (45) exhibited high affinity with a K i value of 20 nM, presenting both in vitro and in vivo activity against T. cruzi. Furthermore, the most promising compounds reduced parasite burden in vivo and showed no toxicity at a dose of 100 mg/kg. These carbamoyl imidazoles are structurally attractive, nonpeptidic, and easy to prepare and synthetically modify. Finally, these results further advance our understanding of the noncovalent mode of inhibition of this pharmaceutically relevant enzyme, building strong foundations for drug discovery efforts.

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