Caracterização estrutural e potencial biotecnológico de duas lectinas pertencentes a família de lectinas homologas a Oscillatoria agardhii agglutinin OAAH

Abstract

Lectins are ubiquitous proteins that has domain(s) for recognition and binding to carbohydrates, being able to interact with various biological fluid molecules and cell surface receptors, presenting a variety of biological effects. Lectins isolated from algae have unique molecular structures, as well as carbohydrate binding specificities and similar biological activities, and are especially interesting for biochemical and biological applications. The lectins isolated from red marine algae Meristiella echinocarpa (MeL) Solieria filiformis (SfL) have purification protocols established and bacteriostatic effect against gram-negative bacteria, antinociceptive, anti-inflammatory and anti-depressive activity for SfL has already been observed. The objective of this work was to determine the primary structures of MeL and SfL, to evaluate their biotechnological potential, to express of recombinant form SfL-1 in E. coli and to determine its three-dimensional structure. The primary structure of a MeL isoform and two SfL isoforms (SfL-1 and SfL- 2) were determined by combination of mass spectrometry and molecular biology techiniques. MeL, SfL-1 and -2 have four internal repeated domains in their amino acid sequences and showed high identity at the carbohydrate binding sites with lectins from the hemagglutinin family of Oscillatoria aghardii (OAAH). The analysis of circular dichroism demonstrated the predominant of β-strands structures with spectra of proteins- βI for both lectins, which presented Tm between 50º and 60 ºC. This lectins MeL and SfL did not exhibit antibacterial activity. MeL was able to agglutinate cells of multidrug resistant Salmonella and V. alginolyticus, and SfL was able to agglutinate E. coli and S. aureus. SfL showed antineoplastic effects against MCF-7 cell line inhibiting 50% cell viability at 125 μg.mL-1 and induced apoptosis via caspase in 24 hours, while the inhibition of fibroblasts was 34% with the same treatment, while MeL stimulated the cytoproliferation of MCF-7 in concentrations up to 250 μg.mL-1. The SfL-1 isoform was produced of recombinant form, exhibited hemagglutinating activity, inhibition for sugars, and secondary structure similar to native. The three-dimensional structure was determined by X-ray diffraction and the lectin is composed of two β-barrel type domains formed by five antiparallel β-strands, which are bound by a short peptide between the β-barrels. In relation to the antibacterial activity, rSfL-1 also showed no activity, but was able to cause a greater bacterial agglutination of the E. coli and S. aureus than the native one. Furtehertmore, rSfL was able to agglutinate V. alginolyticus. Thus, MeL and SfL lectins, which showed differences in bacterial cell recognition, show potential in the initial identification of bacteria in bacterial typing studies. And SfL, which presented antineoplastic effects against the MCF-7 cell line, also has an anticancer potential.