Identificação de proteínas antimicrobianas de flores de alecrim-pimenta (Lippia sidoides): uma nova estratégia no combate a patógenos

Abstract

One of the major global issues in agriculture could be directly related to the severe production crop losses caused by phytopathogenic fungi, especially when infection affects post-harvest cultures. In this view, the fungus Botrytis cinerea is able to cause gray mold in more than 200 species of plants, being considered a major problem for the agribusiness. The use of fungicides is a primary fungi control method in plants, due to velocity and facility of application. However, the continuous use of fungicides may promote the selection of resistant fungi and also the ecosystems contamination. Aiming to find different solutions to this problem, several studies have focused on the search for new alternatives to fungi control, such as plant proteins with antifungal activities (AFPs). In this view, a selection of antimicrobial peptides from flowers, leaves and seeds from Lippia genus and further isolation of antifungal peptides from Lippia sidoides flowers was focused in this work. In this work, the purification and identification of two novel peptides of approximately 10 kDa and 15 kDa from flowers of rosemary-pepper (L. sidoides) was described. L. sidoides flowers were oven dried at 25 oC for 5 days, following protein extraction with a solution containing 0.1% HCl 0.6 M NaCl, and further ammonium sulfate precipitation (100%). After precipitation the extract was dialyzed against distilled water (cut off 1.0 kDa) and lyophilized. The rich fraction was applied onto an Octyl-Sepharose hydrophobic chromatography, followed by HPLC reversed-phase chromatography (Vydac C18-TP). Bioassays using crude extract and in vitro PR fraction indicated the inhibition of Botrytis cinerea growth. N-termini sequences, obtained by the Edman degradation, followed by alignment indicate that these two peptides can be classified as NBS-LRR R proteins. This discovery may help in a near future to the development of biotechnology products such as antifungal drugs and transgenic plants with enhanced resistance to fungal pathogens.