Abstract

BackgroundThe filamentous fungus Aspergillus niger is frequently used for industrial production of fermentative products such as enzymes, proteins and biochemicals. Notable examples of industrially produced A. niger fermentation products are glucoamylase and citric acid. Most notably, the industrial production of citric acid achieves high titers, yield and productivities, a feat that has prompted researchers to propose A. niger to serve as heterologous production host for the industrial production of itaconic acid (IA), a promising sustainable chemical building-block for the fabrication of various synthetic resins, coatings, and polymers. Heterologous production of IA in A. niger has resulted in unexpected levels of metabolic rewiring that has led us to the identification of IA biodegradation pathway in A. niger. In this study we have attempted to identify the final product of the IA biodegradation pathway and analyzed the effect of metabolic rewiring on the bioproduction of 9 industrially relevant organic acids.ResultsIA biodegradation manifests in diminishing titers of IA and the occurrence of an unidentified compound in the HPLC profile. Based on published results on the IA biodegradation pathway, we hypothesized that the final product of IA biodegradation in A. niger may be citramalic acid (CM). Based on detailed HPLC analysis, we concluded that the unidentified compound is indeed CM. Furthermore, by transcriptome analysis we explored the effect of metabolic rewiring on the production of 9 industrially relevant organic acids by transcriptome analysis of IA producing and WT A. niger strains. Interestingly, this analysis led to the identification of a previously unknown biosynthetic cluster that is proposed to be involved in the biosynthesis of CM. Upon overexpression of the putative citramalate synthase and a genomically clustered organic acid transporter, we have observed CM bioproduction by A. niger.ConclusionIn this study, we have shown that the end product of IA biodegradation pathway in A. niger is CM. Knock-out of the IA biodegradation pathway results in the cessation of CM production. Furthermore, in this study we have identified a citramalate biosynthesis pathway, which upon overexpression drives citramalate bioproduction in A. niger.

Highlights

  • The filamentous fungus Aspergillus niger is frequently used for industrial production of fermentative products such as enzymes, proteins and biochemicals

  • To further explore the unexpected level of metabolic rewiring in IA producing A. niger strains, we have analyzed the transcriptome of high and low IA producing strains for genes that are related to biosynthesis and transport of the industrially relevant metabolites citric acid, succinic acid, fumaric acid, malic acid, lactic acid, gluconic acid, oxalic acid, itaconic acid and citraconic acid to see the effects on these genes, which interestingly have led to the identification of another completely unknown citramalic acid (CM) biosynthesis route [13]

  • IA bioconversion in A. niger revisited In our previous communication, we have reported the role of ictA and ichA during IA bioconversion in A. niger [10]

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Summary

Introduction

The filamentous fungus Aspergillus niger is frequently used for industrial production of fermentative products such as enzymes, proteins and biochemicals. Hossain et al Fungal Biol Biotechnol (2019) 6:19 fermentation protocol this resulted in the highest IA titer reported for A. niger (56.5 g/l) [9] This metabolic rewiring towards IA proved more intricate as we have observed induction of genes that are responsible for IA bioconversion and degradation in high IA producing A. niger strains [10]. To further explore the unexpected level of metabolic rewiring in IA producing A. niger strains, we have analyzed the transcriptome of high and low IA producing strains for genes that are related to biosynthesis and transport of the industrially relevant metabolites citric acid, succinic acid, fumaric acid, malic acid, lactic acid, gluconic acid, oxalic acid, itaconic acid and citraconic acid to see the effects on these genes, which interestingly have led to the identification of another completely unknown CM biosynthesis route [13]

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