Orthogonal Assays Clarify the Oxidative Biochemistry of Taxol P450 CYP725A4

ACS Chem Biol. 2016 May 20;11(5):1445-51. doi: 10.1021/acschembio.5b00968. Epub 2016 Mar 16.


Natural product metabolic engineering potentially offers sustainable and affordable access to numerous valuable molecules. However, challenges in characterizing and assembling complex biosynthetic pathways have prevented more rapid progress in this field. The anticancer agent Taxol represents an excellent case study. Assembly of a biosynthetic pathway for Taxol has long been stalled at its first functionalization, putatively an oxygenation performed by the cytochrome P450 CYP725A4, due to confounding characterizations. Here, through combined in vivo (Escherichia coli), in vitro (lipid nanodisc), and metabolite stability assays, we verify the presence and likely cause of this enzyme's inherent promiscuity. Thereby, we remove the possibility that promiscuity simply existed as an artifact of previous metabolic engineering approaches. Further, spontaneous rearrangement and the stabilizing effect of a hydrophobic overlay suggest a potential role for nonenzymatic chemistry in Taxol's biosynthesis. Taken together, this work confirms taxadiene-5α-ol as a primary enzymatic product of CYP725A4 and provides direction for future Taxol metabolic and protein engineering efforts.

MeSH terms

  • Alkenes / chemistry
  • Alkenes / metabolism*
  • Antineoplastic Agents, Phytogenic / chemistry
  • Antineoplastic Agents, Phytogenic / metabolism*
  • Biosynthetic Pathways
  • Cytochrome P-450 Enzyme System / metabolism*
  • Diterpenes / chemistry
  • Diterpenes / metabolism*
  • Escherichia coli / chemistry
  • Escherichia coli / enzymology*
  • Escherichia coli / metabolism
  • Fermentation
  • Metabolic Engineering
  • Models, Molecular
  • Oxidation-Reduction
  • Paclitaxel / chemistry
  • Paclitaxel / metabolism*
  • Substrate Specificity
  • Taxus / chemistry
  • Taxus / enzymology*
  • Taxus / metabolism


  • Alkenes
  • Antineoplastic Agents, Phytogenic
  • Diterpenes
  • taxa-4(5),11(12)diene
  • Cytochrome P-450 Enzyme System
  • Paclitaxel