Estabelecimento da estrutura primária e dados cristalográficos da primeira lectina de Mimosoideae purificada de sementes de Parkia platycephala Benth

Abstract

Most studies on Leguminosae lectins involve members of the Papilionoideae subfamily, while investigations on lectins of the other two subfamilies, Caesalpinoideae and Mimosoideae, are scarce. To date no structural data of any Mimosoideae lectin has been reported. The genus Parkia is regarded as the most primitive group within Leguminosae plants, comprising around 30 species. Parkia platycephala Benth is an important forage tree growing in parts of north-eastern of Brazil. The lectin isolated from seeds of P. platycephala (PPL) belongs to the mannose/glucose-specific group and mannose and oligomannosides with al-3 and 21-6 branch linkages are its best inhibitors. The molecular mass of the purified lectin determined by mass spectrometry was 47 946 ± 6Da (by ESI/MS) and 47 951 ± 9Da ~bv MALDI-TOF/ESI). The apparent molecular mass of the lectin in solutions of pH in the range 4.5-8.5 determined by analytical ultracentrifugation equilibrium sedimentation was 94 ± 3kDa, showing that the protein behaves as a non-pH dependent dimmer. The amino-acid sequence of PPL was determined by Edman degradation of overlapping peptides. This is the first report of the primary structure of a Mimosoideae lectin. The unique cysteine residue of PPL, labeled with eosin-5-iodoacetamide in the absence of denaturing reagents, indicated that the tiol croup is solvent-exposed. The protein contained a blocked N-teeiuinus and a single, nonglvcosylated polypeptide chain composed of three tandemly arranged homologous domains. Each of these domains shares amino-acid sequence similarity with jacalinrelated lectins. Based on this homology, the repeated domain of PPL may display a 13 _rism fold similar to that observed in the crystal structure of the lectin from Helianthus tuberosus. The PPL can be regarded as a mosaic protein composed of two N-terminal Calistegya septum jacalin-like domains (Pkl and Pk2 f3 prism domains) followed by an H.tuberosus jacalin-like domain (Pk3 13 prism domain). We have obtained crystals (2.7 A) that we are trying to solve by molecular replacement using the 13 prism structure of the H tuberosus lectin (PDB entry 1 C3K). The sugar binding specificity of PPL towards mannose, an abundant sugar of surface glycoconjugates of sinuses, bacteria, and fungi, suggests a role for PPL in the defense against plant pathogens.