RESUMO
Hylaseptin-4 (HSP-4, GIGDILKNLAKAAGKAALHAVGESL-NH2) is an antimicrobial peptide originally isolated from Hypsiboas punctatus tree frog. The peptide has been chemically synthetized for structural investigations by CD and NMR spectroscopies. CD experiments reveal the high helical content of HSP-4 in biomimetic media. Interestingly, the aggregation process seems to occur at high peptide concentrations either in aqueous solution or in presence of biomimetic membranes, indicating an increase in the propensity of the peptide for adopting a helical conformation. High-resolution NMR structures determined in presence of DPC-d38 micelles show a highly ordered α-helix from amino acid residues I2 to S24 and a smooth bend near G14. A large separation between hydrophobic and hydrophilic residues occurs up to the A16 residue, from which a shift in the amphipathicity is noticed. Oriented solid-state NMR spectroscopy show a roughly parallel orientation of the helical structure along the POPC lipid bilayer surface, with an insertion of the hydrophobic N-terminus into the bilayer core. Moreover, a noticeable pH dependence of the aggregation process in both aqueous and in biomimetic membrane environments is attributed to a single histidine residue (H19). The protonation degree of the imidazole side-chain might help in modulating the peptide-peptide or peptide-lipid interactions. Finally, molecular dynamics simulations confirm the orientation and preferential helical conformation and in addition, show that HSP-4 tends to self-aggregate in order to stabilize its active conformation in aqueous or phospholipid bilayer environments.
Assuntos
Peptídeos Catiônicos Antimicrobianos/química , Lipossomos/química , Sequência de Aminoácidos , Animais , Peptídeos Catiônicos Antimicrobianos/síntese química , Peptídeos Catiônicos Antimicrobianos/metabolismo , Peptídeos Catiônicos Antimicrobianos/farmacologia , Anuros/metabolismo , Dicroísmo Circular , Escherichia coli/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Lipossomos/metabolismo , Simulação de Dinâmica Molecular , Ressonância Magnética Nuclear Biomolecular , Ligação Proteica , Conformação Proteica em alfa-Hélice , Staphylococcus aureus/efeitos dos fármacosRESUMO
Luehea species are found in almost all Central and South American countries. The present work describes the phytochemical study, isolation, and structural characterisation of friedelin, ß-friedelinol, lupeol, pseudotaraxasterol, ß-sitosterol, betulinic acid, taraxasterol, (-)-epicatechin, ß-sitosterol-3-O-ß-d-glucopyranoside, and (+)-epicatechin-(4ßâ8)-epicatechin from stem barks of Luehea ochrophylla Mart. The structural identification of the isolated compounds was mainly performed by NMR analyses and comparison with the data from literature. These compounds were isolated for the first time in the genus Luehea, except ß-sitosterol glucopyranoside, (-)-epicatechin, and lupeol. Hexane extract (HE) and dichloromethane (DF) and ethyl acetate (AF) fractions exhibited antiparasitic activity against amastigote (intracellular) and trypomastigote culture forms of Trypanosoma cruzi. The ethanol extract (EE), DF, and ethanol fraction (EF) exhibited considerable antifungal activity against Candida albicans. Moreover, extracts and fractions exhibited significant percentage of capture free radicals of 2,2-diphenyl-picrylhydrazyl (DPPH) when compared to the standard of ascorbic acid.
Assuntos
Anti-Infecciosos/farmacologia , Antioxidantes/farmacologia , Antiparasitários/farmacologia , Malvaceae/química , Animais , Anti-Infecciosos/química , Antioxidantes/química , Antiparasitários/química , Chlorocebus aethiops , Avaliação Pré-Clínica de Medicamentos/métodos , Glucosídeos/isolamento & purificação , Glucosídeos/farmacologia , Concentração Inibidora 50 , Espectroscopia de Ressonância Magnética , Triterpenos Pentacíclicos/isolamento & purificação , Triterpenos Pentacíclicos/farmacologia , Casca de Planta/química , Extratos Vegetais/química , Sitosteroides/isolamento & purificação , Sitosteroides/farmacologia , Esteróis/isolamento & purificação , Esteróis/farmacologia , Triterpenos/isolamento & purificação , Triterpenos/farmacologia , Células Vero , Ácido BetulínicoRESUMO
The ethanolic extract from the stem of Brosimum potabile afforded (-)-centrolobin (1), isolated for the first time in this genus. The identification of this compound included COSY and NOESY two-dimensional NMR data.