Análises estruturais de duas lectinas do gênero Centrolobium por cristalografia raio X e bioinformática

Abstract

Lectins, a heterogeneous group of proteins that can bind specifically to free sugars or glycans in a reversible manner, may serve several functions. They are distributed in nature and are widely exploited for their specificity to specific carbohydrates, which are related to cell communication. Furthermore, there is biotechnological potential thanks to the biological activities observed in these proteins. Among them, it is possible to highlight the lectins of the Dalbergieae tribe (Leguminosae family, Papilionoideae subfamily), such as Centrolobium microchaete (CML) and Centrolobium tomentosum (CTL) lectins. However, although there are data on the structural biology of both lectins, the tertiary structure of CML experimentally and the comparison analysis of the interaction between CML and CTL with carbohydrates by bioinformatics have not yet been reported. Thus, the work aims to crystallize and determine the crystallographic structure of CML, and to analyze the interaction of CTM and CML with carbohydrates by bioinformatics. For this, both lectins were purified by affinity chromatography, concentrated and evaluated for purity. Afterwards, crystallization tests were carried out, optimization of the crystallization conditions and obtaining of the tertiary structure after processing the X-ray crystallography data from the National Synchrotron Light Laboratory (LNLS) in Campinas-SP. Subsequently, after data integration, scaling and correction, in addition to refinement with ligands, these structures were the object of in silico experiments using molecular dynamics. As a result, it was possible to obtain the tertiary structure of CML by experimental data, with characterization of the CRD with methyl-mannose-1,3-α-D-mannose (MDM) and analysis of the interactions of CML and CTL residues with MDM by molecular dynamics on a trajectory of 200 ns. Finally, the data showed that these lectins, although specific to mannose, have structural similarities and conservation of several regions of the monomer, such as carbohydrate recognition domain (CRD) and metal binding site (MBS), behave differently without and with the ligand by molecular dynamics, involving variations of polar and non-polar interactions. By this bias, the results will contribute to the field of lectinology, especially with CML structural data and intriguing results on molecular dynamics simulations of lectins with and without ligands. By this bias, the results will contribute to the field of lectinology, especially with CML structural data and intriguing results on molecular dynamics simulations of lectins with and without ligands.