Xanthones are a class of molecules that bind to a number of drug targets and possess a myriad of biological properties. An understanding of xanthone biosynthesis at the genetic level should facilitate engineering of second-generation molecules and increasing production of first-generation compounds. The filamentous fungus Aspergillus nidulans has been found to produce two prenylated xanthones, shamixanthone and emericellin, and we report the discovery of two more, variecoxanthone A and epishamixanthone. Using targeted deletions that we created, we determined that a cluster of 10 genes including a polyketide synthase gene, mdpG, is required for prenyl xanthone biosynthesis. mdpG was shown to be required for the synthesis of the anthraquinone emodin, monodictyphenone, and related compounds, and our data indicate that emodin and monodictyphenone are precursors of prenyl xanthones. Isolation of intermediate compounds from the deletion strains provided valuable clues as to the biosynthetic pathway, but no genes accounting for the prenylations were located within the cluster. To find the genes responsible for prenylation, we identified and deleted seven putative prenyltransferases in the A. nidulans genome. We found that two prenyltransferase genes, distant from the cluster, were necessary for prenyl xanthone synthesis. These genes belong to the fungal indole prenyltransferase family that had previously been shown to be responsible for the prenylation of amino acid derivatives. In addition, another prenyl xanthone biosynthesis gene is proximal to one of the prenyltransferase genes. Our data, in aggregate, allow us to propose a complete biosynthetic pathway for the A. nidulans xanthones.

BackgroundEconomical conversion of lignocellulosic biomass into biofuels and bioproducts is central to the establishment of a robust bioeconomy. This requires a conversion host that is able to both efficiently assimilate the major lignocellulose-derived carbon sources and divert their metabolites toward specific bioproducts.ResultsIn this study, the carotenogenic yeast Rhodosporidium toruloides was examined for its ability to convert lignocellulose into two non-native sesquiterpenes with biofuel (bisabolene) and pharmaceutical (amorphadiene) applications. We found that R. toruloides can efficiently convert a mixture of glucose and xylose from hydrolyzed lignocellulose into these bioproducts, and unlike many conventional production hosts, its growth and productivity were enhanced in lignocellulosic hydrolysates relative to purified substrates. This organism was demonstrated to have superior growth in corn stover hydrolysates prepared by two different pretreatment methods, one using a novel biocompatible ionic liquid (IL) choline α-ketoglutarate, which produced 261 mg/L of bisabolene at bench scale, and the other using an alkaline pretreatment, which produced 680 mg/L of bisabolene in a high-gravity fed-batch bioreactor. Interestingly, R. toruloides was also observed to assimilate p-coumaric acid liberated from acylated grass lignin in the IL hydrolysate, a finding we verified with purified substrates. R. toruloides was also able to consume several additional compounds with aromatic motifs similar to lignin monomers, suggesting that this organism may have the metabolic potential to convert depolymerized lignin streams alongside lignocellulosic sugars.ConclusionsThis study highlights the natural compatibility of R. toruloides with bioprocess conditions relevant to lignocellulosic biorefineries and demonstrates its ability to produce non-native terpenes.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-017-0927-5) contains supplementary material, which is available to authorized users.

Achieving low cost and high efficiency lignocellulose deconstruction is a critical step towards widespread adoption of lignocellulosic biofuels. Certain ionic liquid (IL)-based pretreatment processes effectively reduce recalcitrance of lignocellulose to enzymatic degradation but require either costly separations following pretreatment or novel IL compatible processes to mitigate downstream toxicity. Here we demonstrate at benchtop and pilot bioreactor scales a separation-free, intensified process for IL pretreatment, saccharification, and fermentation of sorghum biomass to produce the sesquiterpene bisabolene, a precursor to the renewable diesel and jet fuel bisabolane. The deconstruction process employs the IL cholinium lysinate ([Ch][Lys]), followed by enzymatic saccharification with the commercial enzyme cocktails Cellic CTec2 and HTec2. Glucose yields above 80% and xylose yields above 60% are observed at all scales tested. Unfiltered hydrolysate is fermented directly by Rhodosporidium toruloides -with glucose, xylose, acetate and lactate fully consumed during fermentation at all scales tested. Bisabolene titers improved with scale from 1.3 g L −1 in 30 mL shake flasks to 2.2 g L −1 in 20 L fermentation. The combined process enables conversion of saccharified IL-pretreated biomass directly to advanced biofuels with no separations or washing, minimal additions to facilitate fermentation, no loss of performance due to IL toxicity, and simplified fuel recovery via phase separation. This study is the first to demonstrate a separation-free IL based process for conversion of biomass to an advanced biofuel and is the first to demonstrate full consumption of glucose, xylose, acetate, and lactic acid in the presence of [Ch][Lys].