Comparative analysis of CYP93E proteins for improved microbial synthesis of plant triterpenoids

Tessa Moses, Johan M Thevelein, Alain Goossens, Jacob Pollier

Research output: Contribution to journalArticlepeer-review

Abstract

Cytochrome P450-dependent monooxygenases (P450s) belonging to the CYP93E subfamily catalyze the C-24 oxidation of the triterpene backbone during the biosynthesis of triterpenoid saponins, which are bioactive plant natural products. In our attempts to produce plant triterpenoids in the yeast Saccharomyces cerevisiae, we observed a poor in vivo catalytic efficiency of the Medicago truncatula CYP93E2. To overcome this biosynthetic bottleneck, we screened publicly available plant genome and transcriptome data for CYP93E subfamily members. Six CYP93E orthologs, exclusively from leguminous plant species, were identified and functionally characterized in S. cerevisiae. Despite the high degree of amino acid conservation, the CYP93E orthologs showed large variations in enzymatic efficiency in yeast. The CYP93E9 from Phaseolus vulgaris showed the highest activity and converted ∼80% of the accumulating in vivo produced substrate β-amyrin to the products olean-12-ene-3β,24-diol and probable 3β-hydroxy olean-12-en-24-oic acid, with a catalytic efficiency that was 61 times higher than that of the M. truncatula CYP93E2. In conclusion, we have expanded the list of functional CYP93E orthologs to a total of nine proteins and show that there are large variations in their catalytic efficiencies when expressed in a heterologous host. Although demonstrated here for the CYP93E family involved in triterpenoid saponin biosynthesis, this phenomenon is undoubtedly extendable to other enzyme families involved in natural product synthesis. Hence, screening for homologous enzymes may become a valuable synthetic biologist’s tool for engineering superior production chassis.
Original languageEnglish
Pages (from-to)47-56
Number of pages10
JournalPhytochemistry
Volume108
Early online date23 Oct 2014
DOIs
Publication statusPublished - 1 Dec 2014

Keywords

  • fabaceae
  • synthetic Biology
  • saccharomyces cerevisiae
  • saponins
  • olean-12-ene-3β,24-diol
  • 3β-Hydroxy olean-12-en-24-oic acid

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