Orthogonal Assays Clarify the Oxidative Biochemistry of Taxol P450 CYP725A4

Bradley Walters Biggs, John Edward Rouck, Amogh Kambalyal, William Arnold, Chin Giaw Lim, Marjan De Mey, Mark Oneil-Johnson, Courtney M. Starks, Aditi Das, Parayil Kumaran Ajikumar

Research output: Contribution to journalArticlepeer-review


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 enzymes 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 Taxols 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.

Original languageEnglish (US)
Pages (from-to)1445-1451
Number of pages7
JournalACS chemical biology
Issue number5
StatePublished - May 20 2016

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine


Dive into the research topics of 'Orthogonal Assays Clarify the Oxidative Biochemistry of Taxol P450 CYP725A4'. Together they form a unique fingerprint.

Cite this