High-efficient production of mushroom polyketide compounds in a platform host Aspergillus oryzae

Abstract

Background Orsellinic acid (2,4-dihydroxy-6-methylbenzoic acid, OA) and its structural analog o-Orsellinaldehyde, have become widely used intermediates in clinical drugs synthesis. Although the research on the biosynthesis of such compounds has made significant progress, due to the lack of suitable hosts, there is still far from the industrial production of such compounds based on synthetic biology. Results With the help of genome mining, we found a polyketide synthase (PKS, HerA) in the genome of the Hericium erinaceus, which shares 60% amino acid sequence homology with ArmB from Armillaria mellea, an identified PKS capable of synthesizing OA. To characterize the function of HerA, we cloned herA and heterologously expressed it in Aspergillus oryzae, and successfully detected the production of OA. Subsequently, the introduction of an incomplete PKS (Pks5) from Ustilago maydis containing only three domains (AMP-ACP-R), which was into herA-containing A. oryzae, the resulted in the production of o-Orsellinaldehyde. Considering the economic value of OA and o-Orsellinaldehyde, we then optimized the yield of these compounds in A. oryzae. The screening showed that when maltose was used as carbon source, the yields of OA and o-Orsellinaldehyde were 57.68 mg/L and 15.71 mg/L respectively, while the yields were 340.41 mg/Kg and 84.79 mg/Kg respectively in rice medium for 10 days. Conclusions Herein, we successfully expressed the genes of basidiomycetes using A. oryzae heterologous host. As a fungus of ascomycetes, which not only correctly splices genes of basidiomycetes containing multiple introns, but also efficiently produces their metabolites. This study highlights that A. oryzae is an excellent host for the heterologous production of fungal natural products, and has the potential to become an efficient chassis for the production of basidiomycete secondary metabolites in synthetic biology. Graphical Abstract

Functional Characterization of Sesquiterpene Synthases and P450 Enzymes in Flammulina velutipes for Biosynthesis of Spiro [4.5] Decane Terpene

“…The sequencing results of RNA extracted from AO-flaX transformants and reverse transcribed into cDNA showed that the introns in these genes were correctly spliced. In previous studies, we used the A. oryzae expression system not only heterologously production diterpene 31−33 and polyketide compounds from basidiomycetes 34 but also identify the functions of 31 STSs from two basidiomycota fungi, C. pseudopinsitus and S. hirsutum, 12 demonstrating that A. oryzae is both an excellent expression host for basidiomycetes and has abundant FPP for the production of various terpenoids.…”

“…In previous studies, we used the A. oryzae expression system not only heterologously production diterpene − and polyketide compounds from basidiomycetes but also identify the functions of 31 STSs from two basidiomycota fungi, C. pseudopinsitus and S.…”

Complexity of fungal polyketide biosynthesis and function

“…For instance, Aspergillus oryzae is a convenient host, because it is a GRAS (generally regarded as safe) organism and produces none or very few own polyketides. It has been used to decipher the alternariol biosynthesis (Wenderoth et al., 2019) and more recently for the production of basidiomycete polyketides (Han et al., 2023). Another host organism is A. nidulans , which has been engineered by deletion of eight entire native BGCs from its genome for a cleaner background and less crosstalk between native and heterologous genes (Chiang et al., 2022).…”

“…For instance, Aspergillus oryzae is a convenient host, because it is a GRAS (generally regarded as safe) organism and produces none or very few own polyketides. It has been used to decipher the alternariol biosynthesis (Wenderoth et al, 2019) and more recently for the production of basidiomycete polyketides (Han et al, 2023).…”

Synthetic Biology Tools for Engineering Aspergillus oryzae

“…Capitalizing on a robust capacity for protein synthesis, secretion, and posttranslational modification, A. oryzae has also shown potential for heterologous protein expression. Recently, the integration of heterologous pathways into the A. oryzae genome through precise genetic manipulation has yielded a diverse array of enzymes and secondary metabolites, exemplified by ergothioneine [5], orsellinic acid [6], chevalone E [7], abscisic acid [8], aphidicolin [9], and others. As genetic manipulation techniques continue to evolve, the utilization of A. oryzae as a host for heterologous protein expression is increasingly maturing.…”