RESUMO
Haemonchus contortus is one of the most economically important parasites infecting small ruminants worldwide. This nematode has shown a great ability to develop resistance to anthelmintic drugs, calling for the development of alternative control approaches. Because lectins recognize and bind to specific carbohydrates and glycan structures present in parasites, they can be considered as an alternative to develop new antiparasitic drugs. Accordingly, this work aimed to investigate the anthelmintic effect of Canavalia brasiliensis (ConBr) lectin against H. contortus and to evaluate a possible interaction of ConBr with glycans of this parasite by molecular docking. ConBr showed significant inhibition of H. contortus larval development with an IC50 of 0.26 mg mL-1. Molecular docking assays revealed that glycans containing the core trimannoside [Man(α1-3)Man(α1-6)Man] of H. contortus interact in the carbohydrate recognition domain of ConBr with an interaction value of MDS = -248.77. Our findings suggest that the inhibition of H. contortus larval development is directly related to the recognition of the core trimannoside present in the glycans of these parasites. This work is the first to report on the structure-function relationships of the anthelmintic activity of plant lectins.
Assuntos
Anti-Helmínticos/química , Anti-Helmínticos/metabolismo , Haemonchus/efeitos dos fármacos , Manosídeos/metabolismo , Lectinas de Plantas/química , Lectinas de Plantas/metabolismo , Animais , Anti-Helmínticos/isolamento & purificação , Sítios de Ligação , Canavalia/química , Haemonchus/crescimento & desenvolvimento , Concentração Inibidora 50 , Larva/efeitos dos fármacos , Larva/crescimento & desenvolvimento , Simulação de Acoplamento Molecular , Lectinas de Plantas/isolamento & purificação , Ligação ProteicaRESUMO
The three-dimensional structure of Dioclea reflexa seed lectin (DrfL) was studied in detail by a combination of X-ray crystallography, molecular docking and molecular dynamics. DrfL was purified by affinity chromatography using Sephadex G-50 matrix. Its primary structure was obtained by mass spectrometry, and crystals belonging to orthorhombic space group P212121 were grown by the vapor diffusion method at 293K. The crystal structure was solved at 1.765Å and was very similar to that of other lectins from the same subtribe. The structure presented Rfactor and Rfree of 21.69% and 24.89%, respectively, with no residues in nonallowed regions of Ramachandran plot. Similar to other Diocleinae lectins, DrfL was capable of relaxing aortic rings via NO induction, with CRD participation, albeit with low intensity (32%). In silico analysis results demonstrated that DrfL could strongly interact with complex N-glycans, components of blood vessel glycoconjugates. Despite the high similarity among Diocleinae lectins, it was also reported that each lectin has unique CRD properties that influence carbohydrate binding, resulting in different biological effects presented by these molecules.
Assuntos
Dioclea/química , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Lectinas de Plantas/química , Lectinas de Plantas/farmacologia , Sementes/química , Sequência de Aminoácidos , Animais , Cristalografia por Raios X , Manosídeos/química , Manosídeos/metabolismo , Lectinas de Plantas/metabolismo , Polissacarídeos/química , Polissacarídeos/metabolismo , Domínios Proteicos , Ratos , Vasodilatadores/química , Vasodilatadores/metabolismo , Vasodilatadores/farmacologiaRESUMO
Plant lectins, especially those purified from species of the Leguminosae family, represent the best-studied group of carbohydrate-binding proteins. Lectins purified from seeds of the Diocleinae subtribe exhibit a high degree of sequence identity notwithstanding that they show very distinct biological activities. Two main factors have been related to this feature: variance in key residues influencing the carbohydrate-binding site geometry and differences in the pH-dependent oligomeric state profile. In this work, we have isolated a lectin from Canavalia boliviana (Cbol) and solved its x-ray crystal structure in the unbound form and in complex with the carbohydrates Man(α1-3)Man(α1-O)Me, Man(α1-4)Man(α1-O)Me and 5-bromo-4-chloro-3-indolyl-α-D-mannose. We evaluated its oligomerization profile at different pH values using Small Angle X-ray Scattering and compared it to that of Concanavalin A. Based on predicted pKa-shifts of amino acids in the subunit interfaces we devised a model for the dimer-tetramer equilibrium phenomena of these proteins. Additionally, we demonstrated Cbol anti-inflammatory properties and further characterized them using in vivo and in vitro models.
Assuntos
Anti-Inflamatórios/farmacologia , Canavalia/química , Edema/tratamento farmacológico , Manosídeos/química , Peritonite/tratamento farmacológico , Lectinas de Plantas/química , Lectinas de Plantas/farmacologia , Sementes/química , Sequência de Aminoácidos , Animais , Sítios de Ligação , Movimento Celular/efeitos dos fármacos , Quimiotaxia/efeitos dos fármacos , Cristalografia por Raios X , Edema/induzido quimicamente , Manosídeos/metabolismo , Modelos Moleculares , Dados de Sequência Molecular , Neutrófilos/citologia , Neutrófilos/efeitos dos fármacos , Peritonite/induzido quimicamente , Conformação Proteica , Ratos , Ratos Wistar , Homologia de Sequência de Aminoácidos , Espectrometria de Massas por Ionização por ElectrosprayRESUMO
Despite their well recognized importance in pathogenesis of Entamoeba histolytica there are few studies dealing with the assembly and secretion of glycoproteins that participate in the adhesion to target cells and in the dissemination of the parasite in infected tissues. Some of these studies refer to the identification and, in some cases, the characterization of glycosyl transferases and glycosidases involved in the biosynthesis of these macromolecules as well as to compartments involved in the amoeba dolichol-linked glycosylation pathway. While an N-glycan trimming alpha-mannosidase has been demonstrated in E. histolytica, little is known on its cellular distribution and properties. Here we describe the presence and partial biochemical characterization of soluble and MMF-associated forms of alpha-mannosidase and the separation of at least three internal membrane structures enriched with this glycosidase. Results are discussed in terms of the possible identity of alpha-mannosidase activity and the potential precursor-product relationship between the two enzyme forms.