RESUMO
Lectins isolated from Canavalia ensiformis (ConA) and Canavalia brasiliensis (ConBr) are promising molecules to prevent cell death. Acute pancreatitis, characterized by acinar cell necrosis and inflammation, presents significant morbidity and mortality. This study has investigated the effects of ConA and ConBr in experimental acute pancreatitis and pancreatic acinar cell death induced by bile acid. Pancreatitis was induced by retrograde pancreatic ductal injection of 3% sodium taurocholate (Na-TC) in male Swiss mice. ConA or ConBr (0.1, 1 or 10 mg/kg) were intravenously applied to mice 1 h and 12 h after induction. After 24 h, the severity of pancreatitis was evaluated by serum amylase and lipase, histopathological changes and myeloperoxidase assay. Pancreatic acinar cells were incubated with ConA (200 µg/ml) or ConBr (200 µg/ml) and taurolithocholic acid 3-sulfate (TLCS; 500 µM). Necrosis and changes in mitochondrial membrane potential (ΔÑ°m) were detected by fluorescence confocal microscopy. Treatment (post-insult) with ConA and ConBr decreased pancreatic damage caused by retrograde injection of Na-TC in mice, reducing pancreatic neutrophil infiltration, edema and necrosis. In addition, ConA and ConBr decreased pancreatic acinar cell necrosis and depolarization of ΔÑ°m caused by TLCS. The inhibition of necrosis was prevented by the lectin domain blockade. In conclusion, ConA and ConBr markedly inhibited in vitro and in vivo damage, effects partly dependent on the interaction with mannose residues on acinar cells. These data support the potential application of these proteins for treatment of acute pancreatitis.
Assuntos
Canavalia , Pancreatite , Doença Aguda , Animais , Anti-Inflamatórios , Canavalia/química , Lectinas/farmacologia , Masculino , Camundongos , Necrose/tratamento farmacológico , Pancreatite/induzido quimicamente , Pancreatite/tratamento farmacológico , Lectinas de Plantas/química , Sementes/químicaRESUMO
OBJECTIVE: To investigate the effect and mechanisms of Andira anthelmia lectin in rat models of acute inflammation. MATERIAL: AAL anti-inflammatory activity was evaluated in Wistar rat models of paw edema and peritonitis. METHODS: AAL (0.01-1 mg/kg i.v.) was injected 30 min before stimulation with carrageenan and with initial and late phase inflammatory mediators into the animals paw or peritoneum for evaluation of cell migration (optical and intravital microscopy), paw edema (plethysmometry and histopathology); hyperalgesia (analgesimetry). RESULTS: AAL inhibited leukocyte migration induced by carrageenan, mainly neutrophils to the peritoneal fluid, decreasing leukocyte adhesion. In the peritoneal fluid, AAL reduced the gene expression of TNF-α and cyclooxygenase, as well the levels of PGE2. AAL inhibited the paw edema induced by carrageenan, serotonin, histamine, TNF-α, PLA2 and PGE2, but not by L-arginine. In this model, AAL also inhibited mechanical hypernociception induced by TNF-α, PGE2, db-cAMP and capsaicin, and the activity of myeloperoxidase in the paw tissues. CONCLUSION: AAL presents anti-inflammatory effect in acute models of rat inflammation involving the participation of prostaglandins, TNF-α and lectin domain.
Assuntos
Fabaceae , Fator de Necrose Tumoral alfa , Animais , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Carragenina , Dinoprostona/metabolismo , Edema/induzido quimicamente , Edema/tratamento farmacológico , Edema/patologia , Fabaceae/metabolismo , Inflamação/patologia , Lectinas , Prostaglandinas , Ratos , Ratos WistarRESUMO
OBJECTIVE AND DESIGN: Sodium channels are highly expressed in nociceptive sensory neurons during hypernociceptive conditions. Based on the presence of a glycosidic portion in the sodium channel ß subunit associated to the antinociceptive effect of leguminous lectins via lectin domain, this study investigated the antinociceptive activity of the lectin isolated from Lonchocarpus araripensis seeds (LAL) in mice behavioral models and in NaV current in the nociceptor of rat dorsal root ganglion (DRG). MATERIAL/METHODS: LAL antinociceptive activity and the participation of opioid system, lectin domain and sodium channels were evaluated in Swiss mice models of nociception (formalin, capsaicin, hot plate, tail flick, von Frey) and in primary cultures of Wistar rats neurons of DRG (patch clamp). RESULTS: LAL presented inhibitory effects in the nociception induced by chemical and mechanical, but not by thermal stimuli and reduced total Na(+) current. LAL activity was inhibited by the lectin association with its binding sugar N-acethyl-glucosamine. CONCLUSION: LAL inhibits peripheral hypernociception by mechanisms that involve the lectin domain, inflammatory mediators and Na(+) channels. The innovative inhibitory action of leguminous lectins on NaV current brings new insights for the investigation of sodium channels role in nociception.
Assuntos
Analgésicos , Fabaceae , Lectinas , Dor/tratamento farmacológico , Canais de Sódio/fisiologia , Analgésicos/farmacologia , Analgésicos/uso terapêutico , Animais , Capsaicina , Formaldeído , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/fisiologia , Temperatura Alta , Lectinas/farmacologia , Lectinas/uso terapêutico , Masculino , Camundongos , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Nociceptividade/efeitos dos fármacos , Estimulação Física , Ratos Wistar , SementesRESUMO
Leishmaniasis is a vector-borne disease transmitted by phlebotomine sand fly. Susceptibility and refractoriness to Leishmania depend on the outcome of multiple interactions that take place within the sand fly gut. Promastigote attachment to sand fly midgut epithelium is essential to avoid being excreted together with the digested blood meal. Promastigote and gut sand fly surface glycans are important ligands in this attachment. The purpose of the present study was to evaluate the interaction of three lectins isolated from leguminous seeds (Diocleinae subtribe), D-glucose and D-mannose-binding, with glycans on Lutzomyia migonei midgut. To study this interaction the lectins were labeled with FITC and a fluorescence assay was performed. The results showed that only Dioclea violacea lectin (DVL) was able to interact with midgut glycans, unlike Cratylia floribunda lectin (CFL) and Canavalia gladiata lectin (CGL). Furthermore, when DVL was blocked with D-mannose the interaction was inhibited. Differences of spatial arrangement of residues and volume of carbohydrate recognition domain (CRD) may be the cause of the fine specificity of DVL for glycans in the surface on Lu. migonei midgut. The findings in this study showed the presence of glycans in the midgut with glucose/mannose residues in its composition and these residues may be important in interaction between Lu. migonei midgut and Leishmania.
Assuntos
Canavalia , Dioclea , Absorção Gastrointestinal/fisiologia , Lectinas de Plantas/metabolismo , Psychodidae , Animais , Lectinas/isolamento & purificação , Lectinas/metabolismo , Lectinas de Plantas/isolamento & purificação , SementesRESUMO
Rhizobium tropici is a Gram-negative bacterium that induces nodules and fixed atmospheric nitrogen in symbiotic association with Phaseolus vulgaris (common bean) and some other leguminous species. Lectins are proteins that specifically bind to carbohydrates and, consequently, modulate different biological functions. In this study, the d-glucose/ d-mannose-binding lectins (from seeds of Dioclea megacarpa, D. rostrata and D. violacea) and D-galactose-binding lectins (from seeds of Bauhinia variegata, Erythina velutina and Vatairea macrocarpa) were purified using chromatographic techniques and evaluated for their effect on the growth of R. tropici CIAT899. All lectins were assayed with a satisfactory degree of purity according to SDS-PAGE analysis, and stimulated bacterial growth; in particular, the Dioclea rostrata lectin was the most active among all tested proteins. As confirmed in the present study, both d-galactose- and d-glucose/d-mannose-binding lectins purified from the seeds of leguminous plants may be powerful biotechnological tools to stimulate the growth of R. tropici CIAT99, thus improving symbiotic interaction between rhizobia and common bean and, hence, the production of this field crop.
Assuntos
Fabaceae/química , Lectinas de Plantas , Rhizobium tropici/crescimento & desenvolvimento , Sementes/química , Lectinas de Plantas/química , Lectinas de Plantas/isolamento & purificação , Lectinas de Plantas/farmacologiaRESUMO
Lectin from the seeds of Dioclea lasiophylla (DlyL) was purified in a single step by affinity chromatography on a Sephadex® G-50 column. DlyL strongly agglutinated rabbit erythrocytes and was inhibited by monosaccharides (D-mannose and α-methyl-D-mannoside) and glycoproteins (ovalbumin and fetuin). Similar to other Diocleinae lectins, DlyL has three chains, α, ß and γ, with mass of 25,569 ± 2, 12,998 ± 1 and 12,588 ± 1 Da, respectively, and has no disulfide bonds. The hemagglutinating activity of DlyL was optimal in pH 8.0, stable at a temperature of 70 °C and decreased in EDTA solution, indicating that lectin activity is dependent on divalent metals. DlyL exhibited low toxicity on Artemia sp. nauplii, but this effect was dependent on the concentration of lectin in solution. DlyL immobilized on cyanogen bromide-activated Sepharose® 4B bound 0.917 mg of ovalbumin per cycle, showing the ability to become a tool for glycoproteomics studies.
Assuntos
Dioclea/química , Hemaglutininas/farmacologia , Lectinas de Ligação a Manose/farmacologia , Extratos Vegetais/farmacologia , Sementes/química , Animais , Artemia , Quelantes/química , Cromatografia de Afinidade , Ácido Edético/química , Eritrócitos/efeitos dos fármacos , Hemaglutinação , Hemaglutininas/química , Hemaglutininas/isolamento & purificação , Concentração de Íons de Hidrogênio , Dose Letal Mediana , Lectinas de Ligação a Manose/química , Lectinas de Ligação a Manose/isolamento & purificação , Ovalbumina/química , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Ligação Proteica , Coelhos , Sefarose/químicaRESUMO
RATIONALE: Lectins are a family of proteins capable of deciphering the glycan code. Several authors have published works about crystallization and mass spectrometry analyses of ConA-like lectins. However, mass spectrometry has never been used to characterize lectin crystal content. In this study, Canavalia grandiflora lectin (ConGF), a ConA-like lectin, was crystallized, part of its primary structure sequenced and the pro-inflammatory activity evaluated. In addition, the crystal content was analyzed by mass spectrometry. METHODS: ConGF was crystallized in the presence of X-Man by hanging-drop vapor diffusion at 293 K and the protein crystal content was analyzed by electrospray ionization in a SYNAPT HDMS mass spectrometer. Partial sequence was obtained by protein digestion with several proteolytic enzymes and the peptides sequenced by liquid chromatography/tandem mass spectrometry (LC/MS/MS). The pro-inflammatory potential of ConGF was also evaluated in the model of rat paw edema. RESULTS: The protein crystals consist of mature α chain and ß and γ fragments measuring 25 612 ± 2 Da, 12 962 ± 2 Da and 12 667 ± 2 Da, respectively. The crystal belongs to the orthorhombic space group I222 (unit cell parameters: a = 67.70, b = 55.90, c = 107.46 Å), assuming a monomer in the asymmetric unit. The solvent content was calculated as 43.50% and the protein content as 2.5 µg. Furthermore, a significant part of the primary structure (65.8%) was determined by mass spectrometry. CONCLUSIONS: As far as we know this is the first report of lectin crystal content characterized by mass spectrometry. Like other ConA-like lectins, GonGF induced paw edema however differing in potency and duration. The observed pro-inflammatory activity suggests that ConGF might be a useful tool in the study of inflammation processes and structure/function relationships.
Assuntos
Edema/induzido quimicamente , Inflamação/induzido quimicamente , Lectinas de Plantas/química , Espectrometria de Massas em Tandem/métodos , Difração de Raios X/métodos , Sequência de Aminoácidos , Animais , Membro Posterior , Dados de Sequência Molecular , Fragmentos de Peptídeos/química , Filogenia , Lectinas de Plantas/farmacologia , Ratos , Ratos Wistar , Sementes/química , Alinhamento de SequênciaRESUMO
Lectins are a structurally heterogeneous group of proteins that have specific binding sites for carbohydrates and glycoconjugates. Because of their biotechnological potential, lectins are widely used in biomedical research. The present study aimed to evaluate the healing potential of the lectin isolated from the marine red alga Bryothamnion seaforthii (BSL). The lectin was purified using ion exchange chromatography with DEAE cellulose and characterized using tandem mass spectrometry. For healing tests, skin wounds were induced in the dorsal thoracic region of mice. These animals were randomly divided into three groups and subjected to topical treatment for 12 days with BSL, bovine serum albumin and 150 mM NaCl. To evaluate the potential of each treatment, the animals were anesthetized and sacrificed on days 2, 7 and 12, respectively. The parameters evaluated included the wound area, the proportion of wound closure and the histological diagnosis. The wound closure was more effective with BSL (Postoperative Day 7 and 12) than controls. The luminal epithelium was completely restructured; the presence of collagen in the dermis and the strongly active presence of young skin annexes demonstrate the potential of treatment with BSL compared with controls. Our findings suggest that BSL has pro-healing properties and can be a potential medical process in the treatment of acute wounds.
Assuntos
Lectinas/química , Lectinas/farmacologia , Rodófitas/química , Cicatrização/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Derme/efeitos dos fármacos , Masculino , Camundongos , Dados de Sequência Molecular , Isoformas de Proteínas , Análise de Sequência de ProteínaRESUMO
Lectins are carbohydrate-binding proteins of non-imune origin. This group of proteins is distributed widely in nature and they have been found in viruses, microorganisms, plants and animals. Lectins of plants have been isolated and characterized according to their chemical, physical-chemical, structural and biological properties. Among their biological activities, we can stress its fungicidal action. It has been previously described the effect of the lectins Dviol, DRL, ConBr and LSL obtained from the seeds of leguminous plants on the growth of yeasts isolated from vaginal secretions. In the present work the experiments were carried out in microtiter plates and the results interpreted by both methods: visual observations and a microplate reader at 530nm. The lectin concentrations varied from 0.5 to 256µg/mL, and the inoculum was established between 65-70% of trammitance. All yeast samples isolated from vaginal secretion were evaluated taxonomically, where were observed macroscopic and microscopic characteristics to each species. The LSL lectin did not demonstrate any antifungal activity to any isolate studied. The other lectins DRL, ConBr and DvioL, showed antifungal potential against yeast isolated from vaginal secretion. These findings offering offer a promising field of investigation to develop new therapeutic strategies against vaginal yeast infections, collaborating to improve women's health.
RESUMO
Surface colonization is an essential step in biofilm development. The ability of oral pathogens to adhere to tooth surfaces is directly linked with the presence of specific molecules at the bacterial surface that can interact with enamel acquired pellicle ligands. In light of this, the aim of this study was to verify inhibitory and antibiofilm action of lectins from the Diocleinaesubtribe against Streptococcus mutans and Streptococcus oralis. The inhibitory action against planctonic cells was assessed using lectins from Canavaliaensi formis (ConA), Canavalia brasiliensis (ConBr), Canavalia maritima (ConM), Canavalia gladiata (CGL) and Canavalia boliviana (ConBol). ConBol, ConBr and ConM showed inhibitory activity on S. mutans growth. All lectins, except ConA, stimulated significantly the growth of S. oralis. To evaluate the effect on biofilm formation, clarified saliva was added to 96-well, flat-bottomed polystyrene plates, followed by the addition of solutions containing 100 or 200 µg/mL of the selected lectins. ConBol, ConM and ConA inhibited the S. mutans biofilms. No effects were found on S. oralis biofilms. Structure/function analysis were carried out using bioinformatics tools. The aperture and deepness of the CRD (Carbohydrate Recognition Domain) permit us to distinguish the two groups of Canavalia lectins in accordance to their actions against S. mutans and S. oralis. The results found provide a basis for encouraging the use of plant lectins as biotechnological tools in ecological control and prevention of caries disease.
Assuntos
Biofilmes/efeitos dos fármacos , Lectinas de Plantas/farmacologia , Streptococcus/efeitos dos fármacos , Streptococcus/crescimento & desenvolvimento , Concanavalina A/farmacologia , Streptococcus mutans/efeitos dos fármacos , Streptococcus mutans/crescimento & desenvolvimento , Streptococcus oralis/efeitos dos fármacos , Streptococcus oralis/crescimento & desenvolvimentoRESUMO
Lectin from the seeds of Dioclea sclerocarpa (DSL) was purified in a single step by affinity chromatography on a Sephadex G-50 column. The primary sequence, as determined by tandem mass spectrometry, revealed a protein with 237 amino acids and 81% of identity with ConA. DSL has a molecular mass of 25,606 Da. The ß and γ chains weigh 12,873 Da and 12,752 Da, respectively. DSL hemagglutinated rabbit erythrocytes (both native and treated with proteolytic enzymes), showing stability even after one hour of exposure to a specific pH range. The hemagglutinating activity of DSL was optimal between pH 6.0 and 8.0, but was inhibited after incubation with D-galactose and D-glucose. The pure protein possesses a molecular mass of 25 kDa by SDS-PAGE and 25,606 Da by mass spectrometry. The secondary structure content was estimated using the software SELCON3. The results indicate that b-sheet secondary structures are predominant in DSL (approximately 42.3% antiparallel b-sheet and 6.7% parallel b-sheet). In addition to the b-sheet, the predicted secondary structure of DSL features 4.1% a-helices, 15.8% turns and 31.3% other contributions. Upon thermal denaturation, evaluated by measuring changes in ellipticity at 218 nm induced by a temperature increase from 20 °C to 98 °C, DSL displayed cooperative sigmoidal behavior with transition midpoint at 84 °C and permitted the observation of two-state model (native and denatured).
Assuntos
Dioclea/química , Lectinas/química , Sementes/química , Espectrometria de Massas em Tandem/métodos , Sequência de Aminoácidos , Animais , Dicroísmo Circular , Dioclea/anatomia & histologia , Hemaglutinação/efeitos dos fármacos , Lectinas/genética , Lectinas/farmacologia , Dados de Sequência Molecular , Peso Molecular , Estrutura Secundária de Proteína , Coelhos , Alinhamento de SequênciaRESUMO
This study investigated the effects of the alga lectin Hypnea cervicornis agglutinin (HCA) on rat zymosan-induced arthritis (ZyA). Zymosan (50-500 µg/25 µL) or sterile saline (Sham) was injected into the tibio-tarsal joint of female Wistar rats (180-200 g). Arthritic animals received morphine (4 mg/kg, intraperitoneal), indomethacin (5 mg/kg, intraperitoneal), or 2% lidocaine (100 µL, subcutaneous). HCA (0.3-3 mg/kg) was administered by intravenous route 30 min before or 2 h after zymosan. 1H-[1,2,4]oxadiazolo[4,3-a]-quinoxalin-1-one (ODQ, 4 µg, intra-articular) was given 30 min prior HCA. Hypernociception was measured every hour until 6 h, time in which animals were sacrificed for evaluation of leukocytes of the intra articular fluid and gene expression of TNF-α, IL-1, IL-10, and iNOS in the joint tissues using PCR techniques. Hypernociception was responsive to morphine and indomethacin, and its threshold was not altered by lidocaine. The post-treatment of HCA reduced both hypernociception and leukocyte influx. This antinociceptive effect was abolished either by ODQ and glibenclamide. HCA also reduced gene expression of iNOS and TNF-α. In conclusion, the antinociceptive effect of HCA in ZyA involves cyclic GMP signalization and selective modulation of cytokine expression.
Assuntos
Artrite/tratamento farmacológico , GMP Cíclico/metabolismo , Citocinas/biossíntese , Lectinas/uso terapêutico , Rodófitas , Zimosan/toxicidade , Analgésicos/isolamento & purificação , Analgésicos/farmacologia , Analgésicos/uso terapêutico , Animais , Artrite/induzido quimicamente , Artrite/metabolismo , Expressão Gênica , Lectinas/isolamento & purificação , Lectinas/farmacologia , Ratos , Ratos Wistar , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologiaRESUMO
Legume lectins have been widely studied and applied for many purposes in the last few decades, but many of their physiological aspects remain elusive. The Diocleinae legume subtribe, which includes intensively explored lectins, such as ConA, presents an unusual and extensive post-translational process which results in minor alterations in protein structure, in turn making its function elusive. Despite previous reports about Diocleinae precursor activity, no structural or functional analyses have ever been carried out to understand the impacts of post-translational processing relative to lectin structure and binding specificity. Here we analyzed the functionality of a non glycosylated, recombinantly expressed lectin precursor from Dioclea grandiflora through inhibition assays, corroborating the experimental data with structural information generated by molecular modeling, docking calculations and molecular dynamics simulations. We demonstrate that Diocleinae precursors are active and share the same carbohydrate specificity as mature lectins. At the same time, however, subtle structural alterations were detected and mostly result in an "incomplete" functionality of the precursor, as consequence of an immature binding site and an unstructured tetramer interface, affecting carbohydrate binding and oligomer formation, respectively.
Assuntos
Dioclea/metabolismo , Lectinas de Plantas/química , Domínios Proteicos , Proteínas Recombinantes/química , Sequência de Aminoácidos , Sítios de Ligação/genética , Carboidratos/química , Biologia Computacional/métodos , Cristalografia por Raios X , Dioclea/genética , Eletroforese em Gel de Poliacrilamida , Escherichia coli/genética , Simulação de Dinâmica Molecular , Lectinas de Plantas/genética , Lectinas de Plantas/metabolismo , Ligação Proteica , Precursores de Proteínas/química , Precursores de Proteínas/genética , Precursores de Proteínas/metabolismo , Proteínas Recombinantes/metabolismo , Análise de Sequência de Proteína , Espectrometria de Massas em TandemRESUMO
A new chromophore-containing agglutinin (Haliclona manglaris agglutinin (HMA)) was isolated from the tropical sponge H. manglaris. HMA was purified by a combination of hydrophobic interaction chromatography and ion exchange chromatography. Native HMA is a heterotrimer formed by two ß-chains (15 kDa) and one α-chain (22 kDa). HMA is a glycoprotein and possesses three intrachain disulfide bonds. Hemagglutinating activity of HMA was stable at neutral pH and temperatures up to 60 °C. HMA was only inhibited by thyroglobulin. Mass spectrometry sequencing and Edman degradation revealed a unique amino acid sequence of about 30%. Moreover, HMA has an organic chromophore of 581 Da, and this characteristic seems to be important to its antioxidant activity. Interestingly, while HMA showed no toxicity against Artemia nauplii and was unable to agglutinate bacterial cells, it did show a high capacity to protect ß-carotene against oxidation. Thus, our findings suggest the putative involvement of HMA in the protection of the sponge against oxidation.
Assuntos
Aglutininas/química , Aglutininas/isolamento & purificação , Corantes Fluorescentes/química , Haliclona/química , Sequência de Aminoácidos , Animais , Antioxidantes/farmacologia , Artemia/efeitos dos fármacos , Carboidratos/análise , Cátions Bivalentes/farmacologia , Cromatografia em Gel , Cromatografia de Fase Reversa , Eletroforese em Gel de Poliacrilamida , Hemaglutinação/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Dados de Sequência Molecular , Peso Molecular , Peptídeos/química , Coelhos , Análise de Sequência de Proteína , Compostos de Sulfidrila/química , Espectrometria de Massas em Tandem , TemperaturaRESUMO
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
Lectins are proteins or glycoproteins with the ability to link to carbohydrates at the cell surface in a specific and reversible manner. Studies have showed that lectins are demonstrate immunomodulatory and antitumor agents. This study aims to evaluate the effect of lectin extracted from the seeds of Canavalia brasiliensis (ConBr) on murine melanoma B16F10 cells by analyzing cell viability, apoptosis index, cell migration, production of cytokines and nitric oxide (NO). Results showed that ConBr was able to reduce cell viability and thwart apoptosis, which could be observed by decrease in cell migration. ConBr also induced NO and IL-12 synthesis. Altogether, these data demonstrate the potential of ConBr as a therapeutic agent for melanoma.
Assuntos
Apoptose/efeitos dos fármacos , Canavalia/imunologia , Melanoma/tratamento farmacológico , Lectinas de Plantas/farmacologia , Animais , Linhagem Celular Tumoral , Movimento Celular/imunologia , Sobrevivência Celular/imunologia , Citocinas/análise , Citometria de Fluxo , Melanoma/imunologia , Camundongos , Óxido Nítrico/análise , Lectinas de Plantas/uso terapêutico , Estatísticas não ParamétricasRESUMO
Canavalia gladiata (CGL), C. maritima (ConM) and C. brasiliensis (ConBr) lectins were evaluated in nociception models. ConBr inhibited first (32%) and second (100%) phases of the formalin test; CGL inhibited only the first (74%) and ConM only the second (59%) phase. Hypernociception evaluated in the Von Frey test was inhibited by ConM (55%), CGL (41%) and ConBr (38%). Acetic acid-induced abdominal writhing was reduced by ConBr (66%), CGL (52%) and ConM (60%). ConBr and CGL effects were reversed by the lectin association with its ligand sugar. The antinociceptive activity of the structural homologous lectins was differentiated by potency, efficacy and mechanisms.
Assuntos
Analgésicos/farmacologia , Lectinas de Plantas/farmacologia , Animais , Masculino , Camundongos , Atividade Motora/efeitos dos fármacosRESUMO
Indole-3-acetic acid (IAA) bound is considered a storage molecule and is inactive. However, some studies have proposed an additional possible regulatory mechanism based on the ability of lectins to form complexes with IAA. We report the first crystal structure of ConM in complex with IAA at 2.15 Å resolution. Based on a tetrameric model of the complex, we hypothesize how the lectin controls the availability of IAA during the early seedling stages, indicating a possible new physiological role for these proteins. A free indole group is also bound to the protein. The ConM interaction with different forms of IAA is a strategy to render the phytohormone unavailable to the cell. Thus, this new physiological role proposed for legume lectins might be a novel mechanism by which IAA levels are decreased in addition to the destruction and formation of new complexes in the later stages of seed germination.
Assuntos
Canavalia/fisiologia , Ácidos Indolacéticos/metabolismo , Lectinas de Plantas/metabolismo , Sementes/metabolismo , Animais , Canavalia/metabolismo , Hemaglutinação/efeitos dos fármacos , Simulação de Acoplamento Molecular , Lectinas de Plantas/química , Lectinas de Plantas/farmacologia , Ligação Proteica , Conformação Proteica , CoelhosRESUMO
Lectins from Diocleinae subtribe belong to the family of legume lectins and are characterized by high identity between their amino acids sequences. It has been shown that punctual differences in amino acid sequences, such as one single amino acid or an alternative conformation, represent changes in biological activities caused by these lectins. Therefore, a more detailed understanding of three-dimensional structures of these proteins is essential for accurate analyzing the relationship between structure and function. In this study lectins purified from the seeds of Dioclea violacea (DVL) and Dioclea rostrata (DRL) were compared with regard to crystal structure and vasorelaxant properties. Differences in structure of lectins were found to be reflected in differences in vasorelaxant effects based on their high specificity and selectivity for cell glycans. Binding activity was related to the position of specific residues in the carbohydrate recognition domain (CRD). DVL complexed structure was solved by X-ray crystallography and was compared to native DVL and DRL. Therefore, DVL was co-crystallized with X-Man, and a molecular modeling with X-Man complexed with DVL was done to compare the complexed and native forms adjusted fit. The relatively narrow and deep CRD in DVL promotes little interaction with carbohydrates; in contrast, the wider and shallower CRD in DRL favors interaction. This seems to explain differences in the level of relaxation induced by DVL (43%) and DRL (96%) in rat aortic rings.
Assuntos
Dioclea/química , Lectinas de Plantas/química , Lectinas de Plantas/farmacologia , Vasodilatadores/química , Vasodilatadores/farmacologia , Sequência de Aminoácidos , Animais , Aorta/efeitos dos fármacos , Aorta/fisiologia , Cristalografia por Raios X , Técnicas In Vitro , Masculino , Manose/química , Manose/metabolismo , Simulação de Acoplamento Molecular , Dados de Sequência Molecular , Lectinas de Plantas/metabolismo , Estrutura Quaternária de Proteína , Estrutura Terciária de Proteína , Ratos , Ratos Wistar , Especificidade da Espécie , Vasodilatadores/metabolismoRESUMO
A new lectin from the marine sponge Haliclona caerulea (H-3) was isolated using a combination of hydrophobic interaction chromatography and ion-exchange chromatography. H-3 is a protein with three distinct bands on SDS-PAGE: 9 kDa, 16 kDa and 18 kDa. Nevertheless, on gel filtration and N-PAGE, H-3 showed a symmetrical peak and a unique band, respectively. Hemagglutinating activity of H-3 was stable at neutral pH and temperatures up to 60 °C. N-Acetylgalactosamine and porcine stomach mucin were the most potent inhibitors of H-3. Primary structure of the lectin was determined using tandem mass spectrometry, and it showed no similarity to any members of the animal lectin families. Top down fragmentation revealed some posttranslational modifications in H-3, including glycosylation. The glycan composition of H-3 was determined, and its structure was predicted. Furthermore, H-3 is a blue protein, binding to a chromophore(-597) by weak interactions, and this is the first time that the interaction between one lectin and a natural chromophore has been shown.