Poster: Secondary Metabolism

Abs # 279: Progress in Taxol biosynthesis: Identification of a taxoid 7b-hydroxylase and a 5a-acetyltransferase

Presenter:	Chau, MyDoanh , mydoanh@hotmail.com
Authors	Chau, MyDoanh  (A) (B)  Croteau, Rodney B. (A) (B)
Affiliations:	(A): Institute of Biological Chemistry (B): Washington State University

Taxol, a highly functionalized diterpene is an important anticancer drug isolated from yew (Taxus) species. The limited supply of this drug from natural sources requires alternative means of Taxol production, which would be carried out officially with cultured Taxus cells. However, improvements in the biological production of Taxol and its semisynthetic precursors require an understanding of the biosynthesis of this natural product and the regulation of the pathway. A multi-step biogenetic scheme has been proposed based on the occurrence of defined taxoid metabolites and on analogy to biosynthetic transformations of simpler terpenoids. Some of these steps include hydroxylations and acylations, which have been shown to be important for the bioactivity of this drug. Recently, two new Taxol biosynthetic enzymes, a taxoid 7b-hydroxylase and an acyltransferase, have been identified. This microsomal cytochrome P450 7b-hydroxylase was functionally expressed in both Saccharomyces cerevesiae and Spodoptera fugiperda/baculovirus systems and shown to convert a surrogate substrate (+)-taxusin to its 7b-hydroxy derivative. The operationally soluble acyltransferase was functionally expressed in Escherichia coli and identified as a 5a-acetyltransferase using taxadien-5a-ol as a substrate. Further experiments using other poly-hydroxylated taxane substrates have demonstrated that this acyltransferase clone differs from a previously-acquired 5a-acetyltransferase in both its specificity and kinetics.