Structural Investigations of Enzymes Responsible for the Modification of Trichothecene Mycotoxins Associated with Fusarium Head Blight

Fusarium head blight (FHB) is a devastating crop disease that primarily affects cereal crops such as wheat, barley, and maize. The infection is caused by several species of fungi from the genus Fusarium and results in a diminished crop yield that is accompanied by the accumulation of trichothecene mycotoxins such as deoxynivalenol, T-2, and nivalenol in the grain. Trichothecene mycotoxins play a critical role in pathogenesis, and can accumulate to threatening levels for both human and animal consumers. The use of trichothecene-modifying enzymes as transgenic resistance factors is being pursued in addition to conventional breeding strategies, which have met with little success to date. Most efforts have been directed at either the trichothecene 3-O-acetyltransferase (TRI101) enzymes from Fusariam spp. or the deoxynivalenol 3-O-glucosyltransferase (DOGT1) enzymes from various plant sources. The choice of enzyme is of critical importance if transgenic resistance strategies are to be successful. Comparatively few of the available trichothecene-modifying enzymes have been tested as transgenic resistance factors. This report aims to bring light to an expanded range of transgenic resistance factors by considering homologous or engineered forms of the currently used enzymes. In vitro and in vivo analyses are presented for several TRI101 orthologs alongside a protein engineering approach to develop an improved TRI101 enzyme. Additionally, a crystal structure for a deoxynivalenol 3-O-glucosyl transferase from Oryza sativa is described. This structure represents the first evidence of key structural features for trichothecene specificity in a broad array of glucosyltransferase enzymes, and will be used to guide future investigations of trichothecene detoxifying enzymes.