Biosynthesis of the highly oxygenated tetracyclic core skeleton of Taxol

Area of Science:

• Biochemistry
• Organic Chemistry
• Molecular Biology

Background:

• Taxol (paclitaxel) is a crucial anticancer drug targeting microtubule dynamics.
• While many enzymes in Taxol biosynthesis are known, the exact pathway from di-oxygenated taxoids to the core skeleton remains unclear.

Purpose of the Study:

• To elucidate the stepwise biosynthetic reactions leading to the Taxol tetracyclic core.
• To identify novel enzymes and mechanisms involved in Taxol biosynthesis.
• To achieve complete biosynthesis of Taxol in engineered yeast.

Main Methods:

• Enzyme characterization and activity assays.
• Confirmation of hydroxylation reaction order.
• Identification of a taxoid 9α-hydroxylase and CYP725A55.
• Identification of an acetyltransferase.
• Metabolic engineering of yeast for Taxol biosynthesis.

Main Results:

• The critical order of the second and third hydroxylation steps was confirmed.
• A novel taxoid 9α-hydroxylase and CYP725A55 (involved in oxetane ester formation) were identified.
• An acetyltransferase responsible for C7-OAc formation was identified.
• The pathway to 1β-dehydroxybaccatin VI was elucidated.
• Complete Taxol biosynthesis was achieved in engineered yeast from taxa-4(20),11(12)-diene-5α-ol.

Conclusions:

• This study significantly advances the understanding of Taxol biosynthesis by elucidating key enzymatic steps and reaction mechanisms.
• The identification of novel enzymes and the successful engineered yeast strain provide a foundation for future Taxol production strategies.