TY - JOUR

T1 - Production of putative diterpene carboxylic acid intermediates of triptolide in yeast

AU - Forman, Victor

AU - Callari, Roberta

AU - Folly, Christophe

AU - Heider, Harald

AU - Hamberger, Björn

PY - 2017

Y1 - 2017

N2 - The development of medical applications exploiting the broad bioactivities of the diterpene therapeutic triptolide from Tripterygium wilfordii is limited by low extraction yields from the native plant. Furthermore, the extraordinarily high structural complexity prevents an economically attractive enantioselective total synthesis. An alternative production route of triptolide through engineered Saccharomyces cerevisiae (yeast) could provide a sustainable source of triptolide. A potential intermediate in the unknown biosynthetic route to triptolide is the diterpene dehydroabietic acid. Here, we report a biosynthetic route to dehydroabietic acid by transient expression of enzymes from T. wilfordii and Sitka spruce (Picea sitchensis) in Nicotiana benthamiana. The combination of diterpene synthases TwTPS9, TwTPS27, and cytochromes P450 PsCYP720B4 yielded dehydroabietic acid and a novel analog, tentatively identified as 'miltiradienic acid'. This biosynthetic pathway was reassembled in a yeast strain engineered for increased yields of the pathway intermediates, the diterpene olefins miltiradiene and dehydroabietadiene. Introduction in that strain of PsCYP720B4 in combination with two alternative NADPH-dependent cytochrome P450 reductases resulted in scalable in vivo production of dehydroabietic acid and its analog from glucose. Approaching future elucidation of the remaining biosynthetic steps to triptolide, our findings may provide an independent platform for testing of additional recombinant candidate genes, and ultimately pave the way to biotechnological production of the high value diterpenoid therapeutic.

AB - The development of medical applications exploiting the broad bioactivities of the diterpene therapeutic triptolide from Tripterygium wilfordii is limited by low extraction yields from the native plant. Furthermore, the extraordinarily high structural complexity prevents an economically attractive enantioselective total synthesis. An alternative production route of triptolide through engineered Saccharomyces cerevisiae (yeast) could provide a sustainable source of triptolide. A potential intermediate in the unknown biosynthetic route to triptolide is the diterpene dehydroabietic acid. Here, we report a biosynthetic route to dehydroabietic acid by transient expression of enzymes from T. wilfordii and Sitka spruce (Picea sitchensis) in Nicotiana benthamiana. The combination of diterpene synthases TwTPS9, TwTPS27, and cytochromes P450 PsCYP720B4 yielded dehydroabietic acid and a novel analog, tentatively identified as 'miltiradienic acid'. This biosynthetic pathway was reassembled in a yeast strain engineered for increased yields of the pathway intermediates, the diterpene olefins miltiradiene and dehydroabietadiene. Introduction in that strain of PsCYP720B4 in combination with two alternative NADPH-dependent cytochrome P450 reductases resulted in scalable in vivo production of dehydroabietic acid and its analog from glucose. Approaching future elucidation of the remaining biosynthetic steps to triptolide, our findings may provide an independent platform for testing of additional recombinant candidate genes, and ultimately pave the way to biotechnological production of the high value diterpenoid therapeutic.

KW - Journal Article

U2 - 10.3390/molecules22060981

DO - 10.3390/molecules22060981

M3 - Journal article

C2 - 28608823

VL - 22

JO - Molecules (Print Archive Edition)

JF - Molecules (Print Archive Edition)

SN - 1431-5157

IS - 6

ER -