ABSTRACT
The Arachis genus belongs to the Dalbergieae tribe, a group of plants that show promising potential novel lectins. Three lectins of the well-known Arachis hypogaea have already been purified, while lectins from related species are still unknown. Genomes of two closely related species, Arachis duranensis and Arachis ipaensis, were recently sequenced. Therefore, this study aimed to establish the three-dimensional structure of Arachis duranensis lectin (ADL) and Arachis ipaensis lectin (AIL) by homology modeling, test their activity against mannosides, and perform molecular dynamics (MD) simulations on these two proteins, both unligated and interacting with mannose or α-methyl-D-mannoside. The fold obtained for the molecular models agrees with data obtained from previous leguminous lectins, showing a conserved jelly roll motif. Docking scores indicate that these lectins have different theoretical binding energy with monosaccharides, disaccharides, and high-mannose glycans. MD simulations revealed that these proteins are α-methyl-D-mannoside-specific, having less stable interactions with mannose. This study thus serves as a guide for further research on lectins of the Arachis genus.
ABSTRACT
Lectins represent a class of proteins or glycoproteins capable of reversibly binding to carbohydrates. Seed lectins from the Dalbergieae tribe (Leguminosae) have structural variability, carbohydrate specificity, and biological effects, such as inflammation, vasorelaxation and cancer antigen binding. To comprehensively address these factors, the present work aimed to establish and characterize the three-dimensional structure of Centrolobium microchaete lectin (CML) by homology modeling, investigate protein-carbohydrate interactions and evaluate its inflammatory effect on mice. Molecular docking was performed to analyze interactions of the lectin with monosaccharides, disaccharides and N-glycans. Two dimannosides, methyl mannose-1,3-α-D-mannose (MDM) and mannose-1,3-α-D-mannose (M13), were used in molecular dynamics (MD) simulations to study the behavior of the carbohydrate-recognition domain (CRD) over time. Results showed an expanded domain within which hydrophobic interactions with the methyl group in the MDM molecule were established, thus revealing novel interactions for mannose-specific Dalbergieae lectins. To examine its biological activities, CML was purified in a single step by affinity chromatography on Sepharose-mannose matrix. The lectin demonstrated inflammatory response in the paw edema model and stimulated leukocyte migration to the animal peritoneal cavities, an effect elicited by CRD. For the first time, this work reports the molecular dynamics of a lectin from the Dalbergieae tribe.
Subject(s)
Fabaceae/chemistry , Molecular Docking Simulation , Molecular Dynamics Simulation , Molecular Structure , Plant Lectins/chemistry , Seeds/chemistry , Animals , Anti-Inflammatory Agents, Non-Steroidal/chemistry , Anti-Inflammatory Agents, Non-Steroidal/isolation & purification , Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Binding Sites , Disease Models, Animal , Edema/drug therapy , Edema/etiology , Edema/pathology , Metals/chemistry , Mice , Plant Lectins/isolation & purification , Plant Lectins/pharmacology , Protein Binding , Protein Interaction Domains and Motifs , Structure-Activity RelationshipSubject(s)
Dioclea/chemistry , Mannose-Binding Lectins/chemistry , Plant Lectins/chemistry , Seeds/chemistry , Animals , Apoptosis/drug effects , Binding Sites , Cell Line, Tumor , Cell Movement/drug effects , Cell Survival/drug effects , Crystallization , Glioma , Mannose-Binding Lectins/isolation & purification , Mannose-Binding Lectins/pharmacology , Molecular Docking Simulation , Plant Lectins/isolation & purification , Plant Lectins/pharmacology , Protein Binding , Protein Conformation , RatsABSTRACT
A native lectin (nPELa), purified from seeds of the species Platypodium elegans, Dalbergieae tribe, was crystallized and structurally characterized by X-ray diffraction crystallography and bioinformatics tools. The obtained crystals diffracted to 1.6Å resolution, and nPELa structure were solved through molecular substitution. In addition, nPELa has a metal binding site and a conserved carbohydrate recognition domain (CRD) similar to other Dalbergieae tribe lectins, such as PAL (Pterocarpus angolensis) and CTL (Centrolobium tomentosum). Molecular docking analysis indicated high affinity of this lectin for different mannosides, mainly trimannosides, formed by α-1,3 or α-1,6 glycosidic bond, as evidenced by the obtained scores. In addition, molecular dynamics simulations were performed to demonstrate the structural behavior of nPELa in aqueous solution. In solution, nPELa was highly stable, and structural modifications in its carbohydrate recognition site allowed interaction between the lectin and the different ligands. Different modifications were observed during simulations for each one of the glycans, which included different hydrogen bonds and hydrophobic interactions through changes in the relevant residues. In addition, nPELa was evaluated for its nociceptive activity in mice and was reported to be the first lectin of the Dalbergieae tribe to show CRD-dependent hypernociceptive activity.
Subject(s)
Fabaceae/chemistry , Nociceptive Pain/drug therapy , Plant Lectins/chemistry , Polysaccharides/chemistry , Animals , Binding Sites , Computational Biology , Crystallography, X-Ray , Hydrogen Bonding , Mannosides/chemistry , Mice , Molecular Docking Simulation , Molecular Dynamics Simulation , Molecular Structure , Nociceptive Pain/pathology , Plant Lectins/administration & dosage , Seeds/chemistryABSTRACT
The lectin from Platypodium elegans seeds (PELa) was purified by affinity chromatography in a mannose-agarose column. The lectin agglutinated rabbit erythrocytes and the agglutinating effect was inhibited by previous incubation with the glycoprotein fetuin, along with N-acetyl-d-glucosamine, D-mannose and its derivatives. The lectin maintained complete activity in temperatures ranging from 40 to 60°C and pH values ranging from 9 to 10. As a glycoprotein, PELa has a carbohydrate content of 2.2%, and its activity requires divalent cations such as Ca2+ and Mn2+. Based on SDS-PAGE, PELa displays a profile similar to that of other Dalbergieae lectins with the main chain of molecular mass around 30kDa and two subunits of 19kDa and 10 kDa each. Two-dimensional (2D) electrophoresis revealed the presence of isoforms with different isoelectric points, and high-performance size exclusion chromatography (HPSEC) was performed to confirm the purity of the sample. The lectin was immobilized in CNBr-activated Sepharose 4B and successfully captured fetuin in solution, demonstrating that this lectin remains active and capable of binding carbohydrates. PELa showed effects different from those of its recombinant form in both pro- and anti-inflammatory tests.
