A novel vasorelaxant lectin purified from seeds of Clathrotropis nitida: Partial characterization and immobilization in chitosan beads

A novel lectin from seeds of Clathrotropis nitida (CNA) was purified and characterized. CNA is a glycoprotein containing approximately 3.3% carbohydrates in its structure. CNA promoted intense agglutination of rabbit erythrocytes, which was inhibited by galactosides and porcine stomach mucin (PSM). The lectin maintained its hemagglutinating activity after incubation in a wide range of temperatures (30-60 °C) and pH (6.0-7.0), and its binding activity was dependent on divalent cations (Ca+2 and Mg+2). SDS-PAGE showed an electrophoretic profile consisting of a single band of 28 kDa, as confirmed by electrospray ionization mass spectrometry, which indicated an average molecular mass of 27,406 ± 2 Da and the possible presence of isoforms and glycoforms. In addition, CNA exhibited no toxicity to Artemia sp. nauplii and elicited reversible and dose-dependent vasorelaxation in precontracted aortic rings. CNA was successfully immobilized on chitosan beads and was able to capture PSM in solution. This study demonstrated that CNA is a lectin that has potential as a biotechnological tool in glycomics and glycoproteomics applications. © 2015 Elsevier Inc

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

Contribution of the carbohydrate-binding ability of Vatairea guianensis lectin to induce edematogenic activity

Vatairea guianensis lectin (VGL), Dalbergiae tribe, is a N-acetyl-galactosamine (GalNAc)/Galactose (Gal) lectin previously purified and characterized. In this work, we report its structural features, obtained from bioinformatics tools, and its inflammatory effect, obtained from a rat paw edema model. The VGL model was obtained by homology with the lectin of Vatairea macrocarpa (VML) as template, and we used it to demonstrate the common characteristics of legume lectins, such as the jellyroll motif and presence of a metal-binding site in the vicinity of the carbohydrate-recognition domain (CRD). Protein-ligand docking revealed favorable interactions with N-acetyl-D-galactosamine, D-galactose and related sugars as well as several biologically relevant N- and O-glycans. In vivo testing of paw edema revealed that VGL induces edematogenic effect involving prostaglandins, interleukins and VGL CRD. Taken together, these data corroborate with previous reports showing that VGL interacts with N- and/or O-glycans of molecular targets, particularly in those presenting galactosides in their structure, contributing to the lectin inflammatory effect. © 2017 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM

Gastrointestinal dysmotility in 5-fluorouracil-induced intestinal mucositis outlasts inflammatory process resolution

Aim To evaluate gastrointestinal motility during 5-fluorouracil (5-FU)-induced intestinal mucositis. Materials and methods Wistar rats received 5-FU (150 mg kg(-1), i.p.) or saline. After the 1st, 3rd, 5th, 15th and 30th day, sections of duodenum, jejunum and ileum were removed for assessment of epithelial damage, apoptotic and mitotic indexes, MPO activity and GSH concentration. In order to study gastrointestinal motility, on the 3rd or 15th day after 5-FU treatment, gastric emptying in vivo was measured by scintilographic method, and stomach or duodenal smooth muscle contractions induced by CCh were evaluated in vitro. Results On the third day of treatment, 5-FU induced a significant villi shortening, an increase in crypt depth and intestinal MPO activity and a decrease in villus/crypt ratio and GSH concentration. On the first day after 5-FU there was an increase in the apoptosis index and a decrease in the mitosis index in all intestinal segments. After the 15th day of 5-FU treatment, a complete reversion of all these parameters was observed. There was a delay in gastric emptying in vivo and a significant increase in gastric fundus and duodenum smooth muscle contraction, after both the 3rd and 15th day. Conclusions 5-FU-induced gastrointestinal dysmotility outlasts intestinal mucositis.CNPq (Brazil

CRLI induces vascular smooth muscle relaxation and suggests a dual mechanism of eNOS activation by legume lectins via muscarinic receptors and shear stress

Lectins are proteins able to recognize carbohydrates, without modifying their structure, via the carbohydrate-recognition domain (CRD). Here, the three-dimensional structure of the mannose-binding lectin isolated from Cymbosema roseum (CRLI) was determined with X-man molecule modeled into the carbohydrate recognition domain. CRLI relaxant activity in thoracic rat aorta was also investigated, and based on the results, a molecular docking of CRLI with heparan sulfate was performed to investigate the possible interaction with mechanoreceptors involved in vasorelaxation. CRLI (IC50 = 12.4 μg mL-1) elicited vasorelaxant response (96%) in endothelialized rat aorta contracted with phenylephrine. Endothelium-derived relaxant factors, extracellular calcium (Ca2+e) and muscarinic receptors were also evaluated as putative participants in the CRLI relaxant effect. CRLI relaxant effect was blocked by L-NAME, a nonselective inhibitor of nitric oxide synthase (NOS), and partially inhibited in a calcium-free solution (0Ca) and by atropine, but it remained unchanged in the presence of indomethacin and TEA. In summary, our data suggest interaction between CRLI and muscarinic receptors located in vascular endothelial cells leading to NOS activation triggered by a mechanism that involves Ca2+e along with the ability of CRLI to interact with heparan sulfate, a highly rated mechanoreceptor involved in eNOS activation. © 2014 Published by Elsevier Inc

Exploring the carbohydrate-binding ability of Canavalia bonariensis lectin in inflammation models

Lectins are a group of proteins of non-immune origin recognized for their ability to bind reversibly to carbohydrates. Researchers have been intrigued by oligosaccharides and glycoconjugates for their involvement as mediators of complex cellular events and then many biotechnological applications of lectins are based on glycocode decoding and their activities. Here, we report a structural and biological study of a ConA-like mannose/glucose-specific lectin fromCanavalia bonariensisseeds, CaBo. More specifically, we evaluate the binding of CaBo with alpha-methyl-D-mannoside (MMA) and mannose-1,3-alpha-D-mannose (M13) and the resultantin vivoeffects on a rat model of acute inflammation. A virtual screening was also carried out to cover a larger number of possible bindings of CaBo.In silicoanalysis demonstrated the stability of CaBo interaction with mannose-type ligands, and the lectin was able to induce acute inflammation in rats with the participation of the carbohydrate recognition domain (CRD) and histamine release. These results confirm the ability of CaBo to interact with hybrid and high-mannoseN-glycans, supporting the hypothesis that CaBo's biological activity occurs primarily through its interaction with cell surface glycosylated receptors