The crystal structure of Canavalia brasiliensis lectin suggests a correlation between its quaternary conformation and its distinct biological properties from Concanavalin A

AbstractCanavalia brasiliensis lectin was isolated from the seeds of a Brazilian autochthonous Leguminosae plant. Despite extensive amino acid sequence similarity with Concanavalin A, C. brasiliensis lectin exerts in vitro and in vivo cellular effects that are markedly different from those displayed by Concanavalin A. We have solved the crystal structure of the C. brasiliensis lectin at 3.0 Å resolution. The three-dimensional structure of the lectin monomer can be superimposed onto that of Concanavalin A with a root-mean-square deviation for all Cα atoms of 0.65 Å. However, this parameter is 0.84 and 1.62 Å when the C. brasiliensis lectin dimer and tetramer, respectively, are compared with the same structures of Concanavalin A. We suggest that these differences in quaternary structure may account for the different biological properties of these two highly related Leguminosae lectins.© 1997 Federation of European Biochemical Societies

Lectins from the Red Marine Algal Species Bryothamnion seaforthii and Bryothamnion triquetrum as Tools to Differentiate Human Colon Carcinoma Cells

The carbohydrate-binding activity of the algal lectins from the closely related red marine algal species Bryothamnion triquetrum (BTL) and Bryothamnion seaforthii (BSL) was used to differentiate human colon carcinoma cell variants with respect to their cell membrane glyco-receptors. These lectins interacted with the cells tested in a dose-dependent manner. Moreover, the fluorescence spectra of both lectins clearly differentiated the cells used as shown by FACS profiles. Furthermore, as observed by confocal microscopy, BTL and BSL bound to cell surface glycoproteins underwent intense internalization, which makes them possible tools in targeting strategies

Morphology, ultrastructure and immunocytochemistry of Hypnea cervicornis and Hypnea musciformis-(Hypneaceae, Rhodophyta) from the coastal waters of Ceará, Brazil

AbstractBased on their morphological and physiological features, red algae comprise a complex and variable group of multiple genera, including Hypnea. In particular, the genus Hypnea J.V. Lamouroux (Cystocloniaceae, Rhodophyta) consists of approximately 54 species, including Hypnea cervicornis and H. musciformis. Lectins were described for both species; however, the localization of these proteins is still unclear. Therefore, this work aimed to characterize the morphology and ultrastructure of Hypnea cervicornis and H. musciformis, as well as localize their lectins at the subcellular level. Samples were collected at Praia do Pacheco (Fortaleza-CE) and processed for light, scanning and transmission electron microscopy, in addition to immunocytochemistry. The studied species presented cortical cell layers, subcortical cells and medullary cells. Based on ultrastructural analysis, these species presented vacuolated cortical cells, with a dense cytoplasm containing chloroplasts. The cell wall consisted of concentric microfibrils embedded in an amorphous matrix. Immunochemistry analysis showed the expression of lectins in the cytoplasm and cell walls. While the structure of the studied algae was similar to the description of other species of the genera under different conditions, this is the first record of algae lectin localization

Modulation of the pharmacological effects of enzymatically-active PLA2 by BTL-2, an isolectin isolated from the Bryothamnion triquetrum red alga

<p>Abstract</p> <p>Background</p> <p>An interaction between lectins from marine algae and PLA₂from rattlesnake was suggested some years ago. We, herein, studied the effects elicited by a small isolectin (BTL-2), isolated from <it>Bryothamnion triquetrum</it>, on the pharmacological and biological activities of a PLA₂isolated from rattlesnake venom (<it>Crotalus durissus cascavella</it>), to better understand the enzymatic and pharmacological mechanisms of the PLA₂and its complex.</p> <p>Results</p> <p>This PLA₂consisted of 122 amino acids (approximate molecular mass of 14 kDa), its pI was estimated to be 8.3, and its amino acid sequence shared a high degree of similarity with that of other neurotoxic and enzymatically-active PLA₂s. BTL-2 had a molecular mass estimated in approximately 9 kDa and was characterized as a basic protein. In addition, BTL-2 did not exhibit any enzymatic activity.</p> <p>The PLA₂and BTL-2 formed a stable heterodimer with a molecular mass of approximately 24–26 kDa, estimated by molecular exclusion HPLC. In the presence of BTL-2, we observed a significant increase in PLA₂activity, 23% higher than that of PLA₂alone. BTL-2 demonstrated an inhibition of 98% in the growth of the Gram-positive bacterial strain, <it>Clavibacter michiganensis michiganensis </it>(Cmm), but only 9.8% inhibition of the Gram-negative bacterial strain, <it>Xanthomonas axonopodis </it>pv <it>passiflorae </it>(Xap). PLA₂decreased bacterial growth by 27.3% and 98.5% for Xap and Cmm, respectively, while incubating these two proteins with PLA₂-BTL-2 inhibited their growths by 36.2% for Xap and 98.5% for Cmm.</p> <p>PLA₂significantly induced platelet aggregation in washed platelets, whereas BTL-2 did not induce significant platelet aggregation in any assay. However, BTL-2 significantly inhibited platelet aggregation induced by PLA₂. In addition, PLA₂exhibited strong oedematogenic activity, which was decreased in the presence of BTL-2. BTL-2 alone did not induce oedema and did not decrease or abolish the oedema induced by the 48/80 compound.</p> <p>Conclusion</p> <p>The unexpected results observed for the PLA₂-BTL-2 complex strongly suggest that the pharmacological activity of this PLA₂is not solely dependent on the presence of enzymatic activity, and that other pharmacological regions may also be involved. In addition, we describe for the first time an interaction between two different molecules, which form a stable complex with significant changes in their original biological action. This opens new possibilities for understanding the function and action of crude venom, an extremely complex mixture of different molecules.</p

ConBr lectin modulates MAPKs and Akt pathways and triggers autophagic glioma cell death by a mechanism dependent upon caspase-8 activation

Glioblastoma multiforme is the most aggressive type of glioma, with limited treatment and poor prognosis. Despite some advances over the last decade, validation of novel and selective antiglioma agents remains a challenge in clinical pharmacology. Prior studies have shown that leguminous lectins may exert various biological effects, including antitumor properties. Accordingly, this study aimed to evaluate the mechanisms underlying the antiglioma activity of ConBr, a lectin extracted from the Canavalia brasiliensis seeds. ConBr at lower concentrations inhibited C6 glioma cell migration while higher levels promoted cell death dependent upon carbohydrate recognition domain (CRD) structure. ConBr increased p38MAPK and JNK and decreased ERK1/2 and Akt phosphorylation. Moreover, ConBr inhibited mTORC1 phosphorylation associated with accumulation of autophagic markers, such as acidic vacuoles and LC3 cleavage. Inhibition of early steps of autophagy with 3-methyl-adenine (3-MA) partially protected whereas the later autophagy inhibitor Chloroquine (CQ) had no protective effect upon ConBr cytotoxicity. ConBr also augmented caspase-3 activation without affecting mitochondrial function. Noteworthy, the caspase-8 inhibitor IETF-fmk attenuated ConBr induced autophagy and C6 glioma cell death. Finally, ConBr did not show cytotoxicity against primary astrocytes, suggesting a selective antiglioma activity. In summary, our results indicate that ConBr requires functional CRD lectin domain to exert antiglioma activity, and its cytotoxicity is associated with MAPKs and Akt pathways modulation and autophagy- and caspase-8- dependent cell death.Fil: Wolin, Ingrid A. V.. Universidade Federal de Santa Catarina; BrasilFil: Heinrich, Isabella A.. Universidade Federal de Santa Catarina; BrasilFil: Nascimento, Ana Paula M.. Universidade Federal de Santa Catarina; BrasilFil: Welter, Priscilla G.. Universidade Federal de Santa Catarina; BrasilFil: Sosa, Liliana del Valle. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones en Ciencias de la Salud. Universidad Nacional de Córdoba. Instituto de Investigaciones en Ciencias de la Salud; ArgentinaFil: de Paul, Ana Lucia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones en Ciencias de la Salud. Universidad Nacional de Córdoba. Instituto de Investigaciones en Ciencias de la Salud; ArgentinaFil: Zanotto Filho, Alfeu. Universidade Federal de Santa Catarina; BrasilFil: Nedel, Cláudia Beatriz. Universidade Federal de Santa Catarina; BrasilFil: Lima, Lara Dias. Universidade Estadual do Ceará; BrasilFil: Osterne, Vinicius Jose Silva. Universidade Estadual do Ceará; BrasilFil: Pinto Junior, Vanir Reis. Universidade Estadual do Ceará; BrasilFil: Nascimento, Kyria S.. Universidade Estadual do Ceará; BrasilFil: Cavada, Benildo S.. Universidade Estadual do Ceará; BrasilFil: Leal, Rodrigo B.. Universidade Federal de Santa Catarina; Brasi

Structural basis for both pro- and anti-inflammatory response induced by mannose-specific legume lectin from Cymbosema roseum

Legume lectins, despite high sequence homology, express diverse biological activities that vary in potency and efficacy. In studies reported here, the mannose-specific lectin from Cymbosema roseum (CRLI), which binds N-glycoproteins, shows both pro-inflammatory effects when administered by local injection and anti-inflammatory effects when by systemic injection. Protein sequencing was obtained by Tandem Mass Spectrometry and the crystal structure was solved by X-ray crystallography using a Synchrotron radiation source. Molecular replacement and refinement were performed using CCP4 and the carbohydrate binding properties were described by affinity assays and computational docking. Biological assays were performed in order to evaluate the lectin edematogenic activity. The crystal structure of CRLI was established to a 1.8 Å resolution in order to determine a structural basis for these differing activities. The structure of CRLI is closely homologous to those of other legume lectins at the monomer level and assembles into tetramers as do many of its homologues. The CRLI carbohydrate binding site was predicted by docking with a specific inhibitory trisaccharide. CRLI possesses a hydrophobic pocket for the binding of α-aminobutyric acid and that pocket is occupied in this structure as are the binding sites for calcium and manganese cations characteristic of legume lectins. CRLI route-dependent effects for acute inflammation are related to its carbohydrate binding domain (due to inhibition caused by the presence of α-methyl-mannoside), and are based on comparative analysis with ConA crystal structure. This may be due to carbohydrate binding site design, which differs at Tyr12 and Glu205 position. © 2011 Elsevier Masson SAS. All rights reserved

Structure of a lectin from Canavalia gladiata seeds: new structural insights for old molecules

<p>Abstract</p> <p>Background</p> <p>Lectins are mainly described as simple carbohydrate-binding proteins. Previous studies have tried to identify other binding sites, which possible recognize plant hormones, secondary metabolites, and isolated amino acid residues. We report the crystal structure of a lectin isolated from <it>Canavalia gladiata </it>seeds (CGL), describing a new binding pocket, which may be related to pathogen resistance activity in ConA-like lectins; a site where a non-protein amino-acid, α-aminobutyric acid (Abu), is bound.</p> <p>Results</p> <p>The overall structure of native CGL and complexed with α-methyl-mannoside and Abu have been refined at 2.3 Å and 2.31 Å resolution, respectively. Analysis of the electron density maps of the CGL structure shows clearly the presence of Abu, which was confirmed by mass spectrometry.</p> <p>Conclusion</p> <p>The presence of Abu in a plant lectin structure strongly indicates the ability of lectins on carrying secondary metabolites. Comparison of the amino acids composing the site with other legume lectins revealed that this site is conserved, providing an evidence of the biological relevance of this site. This new action of lectins strengthens their role in defense mechanisms in plants.</p