Engineering Aspergillus oryzae for the Heterologous Expression of a Bacterial Modular Polyketide Synthase

Abstract

Microbial natural products have had phenomenal success in drug discovery and development yet form distinct classes based on the origin of their native producer. Methods that enable metabolic engineers to combine the most useful features of the different classes of natural products may lead to molecules with enhanced biological activities. In this study, we modified the metabolism of the fungus Aspergillus oryzae to enable the synthesis of triketide lactone (TKL), the product of the modular polyketide synthase DEBS1-TE engineered from bacteria. We established (2S)-methylmalonyl-CoA biosynthesis via introducing a propionyl-CoA carboxylase complex (PCC); reassembled the 11.2 kb DEBS1-TE coding region from synthetic codon-optimized gene fragments using yeast recombination; introduced bacterial phosphopantetheinyltransferase SePptII; investigated propionyl-CoA synthesis and degradation pathways; and developed improved delivery of exogenous propionate. Depending on the conditions used titers of TKL ranged from <0.01–7.4 mg/L. In conclusion, we have demonstrated that A. oryzae can be used as an alternative host for the synthesis of polyketides from bacteria, even those that require toxic or non-native substrates. Our metabolically engineered A. oryzae may offer advantages over current heterologous platforms for producing valuable and complex natural products.

Microbial chassis engineering drives heterologous production of complex secondary metabolites

“…The extensive study in the field of synthetic biology tools for driving enhanced gene expression in A. oryzae reported two constitutive promoter sequences grounding on scalable transcriptome datasets, which showed favorable activities comparable to the two previously characterized strong promoters with regard to the ustiloxin B production (Umemura et al 2020). Additionally, the metabolism modification of A. oryzae has phenomenal success in enabling efficient heterologous expression of bacterial modular PKS via rapid and simple metabolic engineering, exemplified as introduction of bacterial propionyl-CoA carboxylase complex (PCC) and PPTase, as with the reassembly of the modular polyketide synthase DEBS1-TE coding region (Feng et al 2021). This work manifested the metabolically engineered A.…”

Curiouser and curiouser: progress in understanding the programming of iterative highly-reducing polyketide synthases

“…Type III systems, analogous to the classical plant chalcone synthases, are known to occur in fungi where they catalyse relatively simple reactions. 1 While modular Type I PKS have been detected in some fungal genomes, 2 and heterologous expression of bacterial modular PKS has been shown to be possible in fungi, 3 the overwhelming majority of fungal PKS are Type I iterative systems. These are classified as non-reducing (nr-PKS), partially reducing (pr-PKS) and highly reducing (hr-PKS) systems.…”

Heterologous Expression of Secondary Metabolite Genes in Trichoderma reesei for Waste Valorization

“…Heterologous hosts include Escherichia coli [4], Saccharomyces cerevisiae [5], Aspergillus oryzae [1] and Aspergillus nidulans [6]. Many common host organisms such as S. cerevisiae and E. coli are conveniently manipulated, but require tight control of fermentation conditions and media components [1,5,[7][8][9][10][11]. However, filamentous fungi offer high potential as hosts for secondary metabolite production as they can grow rapidly on a wide variety of substrates.…”