Formation of ellagic acid precursors in Fragaria

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Strawberry (Fragaria x ananassa) contains high amounts of bioactive polyphenols, in particular ellagic acid/ellagitannins. Recent significant findings strongly suggest that ellagic acid/ellagitannins provide effective protection against diverse human diseases including cardiovascular diseases, neurodegenerative diseases, and hormonal cancers like prostate and breast cancer. Ellagitannins possess strong antimicrobial activity towards bacteria, yeasts and viruses and thus serve as natural defence against diverse range of microbial infections. Although the biological significance of these compounds is widely acknowledged the genetic background of their biosynthetic pathway is almost unknown.We are planning to clone and heterologously express genes coding for putative gallate:UDP-glucose glucosyltransferases (GT) and gallate acyltransferases (AT) assumed to be involved in the formation of ellagitannin precursors. Candidate genes will be identified on the basis of sequence information of biochemically characterized (hydroxyl)benzoate GTs and acyl-glucose dependent ATs in combination with expression analysis performed by next-generation sequencing (NGS) and quantitative PCR. Alternatively, we will isolate and purify ellagitannin-pathway enzymes from F. x ananassa by activity guided assays. The proteins will be sequenced and the corresponding open reading frames extracted from the recently published genome sequence of F. vesca. It is supposed that both Fragaria species share a common ancestor. Recombinant proteins will be expressed in Escherichia coli or Saccharomyces cerevisiae and functionally characterized. The data can be used to develop molecular markers for high ellagic acid/ellagitannin content that will shorten the breeding period towards cultivars with additional health benefits. Besides, the result can be applied for the biotechnological production of ellagic acid derivatives.