Metabolic responses of Aspergillus terreus under low dissolved oxygen and pH levels
- 84 Downloads
The metabolic responses of Aspergillus terreus NRRL1960 to stress conditions (low dissolved oxygen and pH with limited nitrogen and phosphate) in the two-phase fermentation were investigated in this study. The fermentation kinetics suggested that itaconate production was suppressed under low dissolved oxygen (DO) concentrations. A slight change in pH caused a significant change in itaconate production. The transcriptomic data revealed that under low DO concentration, the glycolytic pathway was uncoupled from the oxidative phosphorylation, resulting in the activation of substrate-level phosphorylation as an alternative route for ATP regeneration. The downregulation of pdh genes, the genes encoding ATP synthase and succinate dehydrogenase, confirmed the observation of the uncoupling of the oxidative TCA cycle from glycolysis. It was found that the upregulation of pyc resulted in a large pool of oxaloacetate in the cytosol. This induced the conversion of oxaloacetate to malate. The upregulation of the gene encoding fumarate hydratase with the subsequent formation of fumarate was found to be responsible for the regeneration of NADPH and ATP under the condition of a low dissolved oxygen level. The large pool of oxaloacetate drove itaconic acid production also via the oxidative TCA cycle. Nevertheless, the downregulation of ATP synthase genes resulted in the deficiency of the proton-pumping H+ ATPase and the subsequent stress due to the failure to maintain the physiological pH. This resulted in itaconate production at a low titer. The fermentation kinetics and the transcriptomic data provided in this study can be used for further process optimization and control to improve itaconate production performance.
KeywordsA. terreus Central metabolic pathway Gene expression Metabolite production Fermentation
The authors would like to thank Dr. Kaemwich Jantama for his fruitful discussion on transcriptomic analysis. The work in RNA-seq gene expression analysis performed by D. Ashley Hill and her colleagues at ResourcePath, Sterlin, VA, USA are highly appreciated. The authors also thank Timothy Wesselman and Roshni Patel from OnRamp Bioinformatics, Inc. (San Diego, CA, USA) for their outstanding work on analyzing the RNA-seq data and differential gene expression analysis.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
This article does not contain any studies with human participants or animals performed by any of the authors.
- Birren BW, Lander ES, Galagan JE, et al (2005) Annotation of the Aspergillus terreus NIH264 genome. EMBL/GenBank/DDBJ databasesGoogle Scholar
- Karaffa L, Diaz R, Papp B, Fekete E, Sandor E, Kubicek CP (2015) A deficiency of manganese ions in the presence of high sugar concentrations is the critical parameter for achieving high yields of itaconic acid by Aspergillus terreus. Appl Microbiol Biotechnol 99:7937–7944CrossRefPubMedGoogle Scholar
- Protein knowledgebase (UniProtKB) of Aspergillus terreus NIH2624. Available from www.uniprot.org/uniprot/?query=A+terreus+&sort=score. Accessed March 13, 2017
- Romano AH, Bright MM, Scott WE (1967) Mechanism of fumaric acid accumulation in Rhizopus nigricans. J Biotechnol 93:600–604Google Scholar
- Songserm P, Thitiprasert S, Tolieng V, Piluk J, Tanasupawat S, Assabumrungrat S, Yang ST, Karnchanatat A, Thongchul N (2015) Regulating pyruvate carboxylase in the living culture of Aspergillus terreus NRRL1960 by L-aspartate for enhanced itaconic acid production. Appl Biochem Biotechnol 177:595–609CrossRefPubMedGoogle Scholar
- Thitiprasert S, Pimtong V, Kodama K, Sooksai S, Tanasupawat S, Assabumrungrat S, Tolieng V, Thongchul N (2016) Correlative effect of dissolved oxygen and key enzyme inhibitors responsible for L-lactate production by immobilized Rhizopus oryzae NRRL395 cultivated in a static bed bioreactor. Process Biochem 51:204–212CrossRefGoogle Scholar