Cristalização, estrutura tridimensional e atividade biológica de uma lectina de sementes de Canavalia maritima.

Abstract

Crystals of Canavalia maritima seeds lectin (ConM) were obtained using the vapor diffusion method in hanging drop by sparse matrix with reservatory solution containing 0,1 M HEPES, pH 8,43, 4% Polyethylene Glycol 400 and 2 M ammonium sulfate. The soaking technic was used to produce lectin crystals complexed with the carbohydrates maltose and trehalose. ConM native crystals diffracted to 1.56 Å resolution, belonging to the primitive orthorhombic space group P21212, with unit-cell parameters a=67.9, b=71.0, c=97.6 Å, consisting in a dimer in the asymmetric unit (VM = 2,4 A3 Da-1), with two metal binding sites per monomer, and loops involved in the molecular oligomerization. The structure was solved by standard molecular replacement techniques. The wrong amino acids residues previously suggested by manual sequencing by Perez et al. (1991) are now determinated as I17, I53, S129, S134, G144, S164, P165, S187, V190, S169, T196 and S202. These modifications confer 98% of similarity between ConM, ConA and ConBr. ConM structure presents a high level of tertiary protein folding, typically of legume lectins. Despite the specificity and the numbers of hydrogen bonds formed in the interaction with the disaccharides maltose and trehalose be the same, structurally changes occurs in the pattern of formation of the hydrogen bonds, especially in the distribution in the number of hydrogen bonds between the two glucose residues of the disaccharides. Our biological activity data show that the relaxant concentration-dependent effect of the seed lectin of Canavalia maritima on isolated aortic rings probably occurs via interaction with a specific lectin-binding site on the endothelium, resulting in a release of nitric oxide.