Renewable energy sources play a central role in the sustainability of global resources. In this context, sugarcane derivatives stand out as major contributors to bioenergy production. However, the occurrence of severe phytopathologies, such as leaf scald, affects the productivity of sugarcane crops. Leaf scald is a widespread disease caused by the bacteria Xanthomonas albilineans that has a dramatic impact on crop productivity, including reduced yields and drop in quality of the juice. The absence of chemical or biological agents against leaf scald boosts the research and development of bioactive molecules as effective and selective agrochemicals. Because of that, phosphopantetheinyl transferase (XaPPT), benzoate-coenzyme A ligase (XaBCL) and N5, N10 - methylenetetrahydrofolate dehydrogenase-cyclohydrolase (XaFolD) enzymes were selected as attractive molecular targets for agrochemical design. The XaPPT and XaBCL play key roles on the biosynthesis of albicidins, phytotoxins that block the differentiation of chloroplasts. XaFolD acts on folate metabolism, crucial to biosynthesis of thymidine, purines and amino acids. These enzymes are essential to X. albilineans viability and pathogenicity, therefore they were selected as attractive molecular targets for structural molecular biology studies. The enzymes were successfully cloned and soluble expressed in large scale. The purification by chromatography led to proteins with high degree of purity (>95%). Crystallization assays employing vapor diffusion and sparse matrix techniques identified attractive conditions to XaPPT (50 mM KH2PO4 pH 5.5, 20% PEG 8000 and 10 mM sodium acetate) XaBCL (100 mM sodium cacodylate pH 7.3, 100 mM calcium acetate and 18% PEG 8000) and XaFolD (0.1 mM Tris-HCl pH 8.0, 200 mM C2H2MgO4, and 20% PEG 3350) crystallization. Pure protein is crucial to the development of structural biology and structure-based drug design strategies aiming at the discovery of new agrochemicals. The selected enzymes are promising targets for medicinal chemistry efforts aimed at developing new alternatives for leaf scald control.

Structural molecular biology toward the discovery of new agrochemicals: Applications in sugarcane leaf scald disease

248th National Meeting (2014)

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