Investigation of Fungal Iterative Polyketide Synthase Functions Using Partially Assembled Intermediates

Abstract

Iterative polyketide synthases (PKS) are large multifunctional enzymes that resemble eukaryotic fatty acid synthases, but can make highly functionalized secondary metabolites using complex and unresolved programming rules. During biosynthesis of the kinase inhibitor hypothemycin by Hypomyces subiculosus, a highly-reducing iterative PKS, Hpm8, cooperates with a non-reducing iterative PKS, Hpm3, to construct the advanced intermediate dehydrozearalenol (DHZ). The identity of putative intermediates in the formation of the highly reduced hexaketide portion of DHZ could be confirmed by incorporation of 13C-labeled N-acetylcysteamine (SNAC) thioesters using the purified enzymes. The results show that Hpm8 can accept SNAC thioesters of intermediates ready for transfer from its acyl carrier protein (ACP) domain to its ketosynthase (KS) domain and assemble them into DHZ in cooperation with Hpm3. Addition of certain structurally modified analogs of intermediates to Hpm8 and Hpm3 can produce DHZ derivatives.

A Single Module Type I Polyketide Synthase Directs de Novo Macrolactone Biogenesis during Galbonolide Biosynthesis in Streptomyces galbus

“…1A). This type of divergence in ␤-keto group modification is not known for bacterial iterative PKS systems and has some precedents only in fungal type I PKS systems (10,11). An iterative PKS system that incorporates methylmalonyl-CoA has not been reported, whereas the ability of FAS to incorporate methylmalonyl-CoA has been observed for mycocerosic acid synthase (23).…”

“…Fungal type I PKSs are iterative, but their products are not limited to aromatic compounds (9). The highly reducing, iterative fungal PKSs synthesize complex polyketide structures by selectively employing post-condensational modification activities in a highly programmed manner (10,11). In these fungal PKSs, the selective usage of the modification activities in each condensation step generates complex polyketide structures.…”

Saccharomyces cerevisiae as a tool for mining, studying and engineering fungal polyketide synthases

“…Zhou et al and Gao et al both fed large acyl SNAC (N-acetylcysteamine thioester) substrates to fungal iPKSs expressed or purified from BJ5464-NpgA (Zhou et al, 2010; Gao et al, 2013). Both groups were able to produce unnatural analogs of the relevant natural product.…”

“…Both groups were able to produce unnatural analogs of the relevant natural product. Gao et al found that the efficiency of incorporation of the unnatural precursor analogs depended on the nature of the structural changes between these analogs and the natural precursors (Gao et al, 2013). …”

Fungal polyketide engineering comes of age

“…Further building on the impressive body of work compiled by the Tang and Vederas laboratories on fungal iPKS systems (9,16,17), in the present study, Xu et al take a significant step forward and explore how structural diversity for BDLs can be generated by pairwise combination of hrPKS and nrPKS modules derived from four different BDL biosynthetic gene clusters. In addition to CcRad and AtCur gene loci, they further expand their approach to include the newly discovered lasiodiplodin and resorcylide biosynthetic iPKSs from Lasiodiplodia theobromae (LtLas) and Acremonium zeae (AzRes), respectively (11), to provide 16 possible hrPKS-nrPKS combinations.…”