Publications

• Reconstruction of the biosynthetic pathway for the core fungal polyketide scaffold rubrofusarin in Saccharomyces cerevisiae.

  Rugbjerg P., Naesby M., Mortensen U.H., Frandsen R.J.

  <h4>Background</h4>Fungal polyketides include commercially important pharmaceuticals and food additives, e.g. the cholesterol-lowering statins and the red and orange monascus pigments. Presently, production relies on isolation of the compounds from the natural producers, and systems for heterologo ... >> More

  <h4>Background</h4>Fungal polyketides include commercially important pharmaceuticals and food additives, e.g. the cholesterol-lowering statins and the red and orange monascus pigments. Presently, production relies on isolation of the compounds from the natural producers, and systems for heterologous production in easily fermentable and genetically engineerable organisms, such as Saccharomyces cerevisiae and Escherichia coli are desirable. Rubrofusarin is an orange polyketide pigment that is a common intermediate in many different fungal biosynthetic pathways.<h4>Results</h4>In this study, we established a biosynthetic pathway for rubrofusarin in S. cerevisiae. First, the Fusarium graminearum gene encoding polyketide synthase 12 (PKS12) was heterologously co-expressed with the Aspergillus fumigatus gene encoding phosphopantetheinyl transferase (npgA) resulting in production of YWA1. This aromatic heptaketide intermediate was converted into nor-rubrofusarin upon expression of the dehydratase gene aurZ from the aurofusarin gene cluster of F. graminearum. Final conversion into rubrofusarin was achieved by expression of the O-methyltransferase encoding gene aurJ, also obtained from the aurofusarin gene cluster, resulting in a titer of 1.1 mg/L. Reduced levels of rubrofusarin were detected when expressing PKS12, npgA, and aurJ alone, presumably due to spontaneous conversion of YWA1 to nor-rubrofusarin. However, the co-expression of aurZ resulted in an approx. six-fold increase in rubrofusarin production.<h4>Conclusions</h4>The reconstructed pathway for rubrofusarin in S. cerevisiae allows the production of a core scaffold molecule with a branch-point role in several fungal polyketide pathways, thus paving the way for production of further natural pigments and bioactive molecules. Furthermore, the reconstruction verifies the suggested pathway, and as such, it is the first example of utilizing a synthetic biological "bottom up" approach for the validation of a complex fungal polyketide pathway. << Less

  Microb. Cell Fact. 12:31-31(2013) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• Two novel classes of enzymes are required for the biosynthesis of aurofusarin in Fusarium graminearum.

  Frandsen R.J., Schuett C., Lund B.W., Staerk D., Nielsen J., Olsson S., Giese H.

  Previous studies have reported the functional characterization of 9 out of 11 genes found in the gene cluster responsible for biosynthesis of the polyketide pigment aurofusarin in Fusarium graminearum. Here we reanalyze the function of a putative aurofusarin pump (AurT) and the two remaining orpha ... >> More

  Previous studies have reported the functional characterization of 9 out of 11 genes found in the gene cluster responsible for biosynthesis of the polyketide pigment aurofusarin in Fusarium graminearum. Here we reanalyze the function of a putative aurofusarin pump (AurT) and the two remaining orphan genes, aurZ and aurS. Targeted gene replacement of aurZ resulted in the discovery that the compound YWA1, rather than nor-rubrofusarin, is the primary product of F. graminearum polyketide synthase 12 (FgPKS12). AurZ is the first representative of a novel class of dehydratases that act on hydroxylated γ-pyrones. Replacement of the aurS gene resulted in accumulation of rubrofusarin, an intermediate that also accumulates when the GIP1, aurF, or aurO genes in the aurofusarin cluster are deleted. Based on the shared phenotype and predicted subcellular localization, we propose that AurS is a member of an extracellular enzyme complex (GIP1-AurF-AurO-AurS) responsible for converting rubrofusarin into aurofusarin. This implies that rubrofusarin, rather than aurofusarin, is pumped across the plasma membrane. Replacement of the putative aurofusarin pump aurT increased the rubrofusarin-to-aurofusarin ratio, supporting that rubrofusarin is normally pumped across the plasma membrane. These results provide functional information on two novel classes of proteins and their contribution to polyketide pigment biosynthesis. << Less

  J. Biol. Chem. 286:10419-10428(2011) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.