Biosynthesis and genetic control of the raspberry aroma

We have been studying the biosynthesis of the raspberry ketone in raspberries and their tissue cultures. We have purified the first enzyme of the pathway,benzalacetone synthase and are investigating presently its genetic control. Our ultimate goal is to introduce multiple copies of the benzalacetone synthase gene into raspberries to increase aroma production. To elucidate the genetic control of the benzalacetone synthase, we chose a genomic approach that is summarized here. Three polyketide synthase (PKS) genes have been cloned, sequenced and characterized from cell suspension cultures of raspberry (Rubus idaeus L. cv Royalty). The three genes show high similarity in both their nucleotide and amino acid sequence. RiPKS1 and RiPKS3 show four nucleotide differences resulting in four amino acid exchanges, while RiPKS2 has three additional differences at the nucleotide level resulting in two additional amino acid exchanges between RiPKS1 and RiPKS2. Enzymological analysis of the proteins expressed in E. coli showed that RiPKS1 is a typical naringenin chalcone sythase (CHS) that produced narigenin as the main reaction product, and p-coumaryltriacetic acid lactone (CTAL) as a minor by-product under in vitro conditions. RiPKS2 did not use any of the aromatic CoA derivatives tried, and no enzyme activity of this protein could be detected. RiPKS3 is an aromatic PKS with limited cyclization capacity whose major reaction product is CTAL. Narigenin was a minor reaction product under the same in vitro reaction conditions. See Zheng et al, (2001) Plant Molec.Biol. 51:1-15. This is a fun project that we have done in collaboration with the Schröders in Freiburg, Germany (see their website: http://www.biologie.uni-freiburg.de/data/schroeder/schroeder.html).