Applied Microbiology and Biotechnology

, Volume 100, Issue 17, pp 7541–7548 | Cite as

Identification of an itaconic acid degrading pathway in itaconic acid producing Aspergillus terreus

  • Mei Chen
  • Xuenian Huang
  • Chengwei Zhong
  • Jianjun Li
  • Xuefeng LuEmail author
Biotechnologically relevant enzymes and proteins


Itaconic acid, one of the most promising and flexible bio-based chemicals, is mainly produced by Aspergillus terreus. Previous studies to improve itaconic acid production in A. terreus through metabolic engineering were mainly focused on its biosynthesis pathway, while the itaconic acid-degrading pathway has largely been ignored. In this study, we used transcriptomic, proteomic, bioinformatic, and in vitro enzymatic analyses to identify three key enzymes, itaconyl-CoA transferase (IctA), itaconyl-CoA hydratase (IchA), and citramalyl-CoA lyase (CclA), that are involved in the catabolic pathway of itaconic acid in A. terreus. In the itaconic acid catabolic pathway in A. terreus, itaconic acid is first converted by IctA into itaconyl-CoA with succinyl-CoA as the CoA donor, and then itaconyl-CoA is hydrated into citramalyl-CoA by IchA. Finally, citramalyl-CoA is cleaved into acetyl-CoA and pyruvate by CclA. Moreover, IctA can also catalyze the reaction between citramalyl-CoA and succinate to generate succinyl-CoA and citramalate. These results, for the first time, identify the three key enzymes, IctA, IchA, and CclA, involved in the itaconic acid degrading pathway in itaconic acid producing A. terreus. The results will facilitate the improvement of itaconic acid production by metabolically engineering the catabolic pathway of itaconic acid in A. terreus.


Itaconic acid Aspergillus terreus Degradation pathway Metabolic engineering 



This work was supported by the National High Technology Research and Development Program of China (2015AA021003), the National Natural Sciences Foundation of China (31500042 and 31400080), the Key Research Program of the Chinese Academy of Sciences (KSZD-EW-Z-016-2), and the Science and Technology Service Network Initiative of the Chinese Academy of Sciences (KFJ-EW-STS077-RW10).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.

Ethical approval

This article does not contain any studies with human participants or animals.

Supplementary material

253_2016_7554_MOESM1_ESM.pdf (819 kb)
ESM 1 (PDF 819 kb)


  1. Adler J, Wang SF, Lardy HA (1957) The metabolism of itaconic acid by liver mitochondria. J Biol Chem 229(2):865–879PubMedGoogle Scholar
  2. Arpai J (1958) Itaconicoxidase: an enzyme from an ultra-violet-induced mutant of Aspergillus terreus. Nature 182(4636):661–662. doi: 10.1038/182661a0 CrossRefPubMedGoogle Scholar
  3. Blumhoff ML, Steiger MG, Mattanovich D, Sauer M (2013) Targeting enzymes to the right compartment: metabolic engineering for itaconic acid production by Aspergillus niger. Metab Eng 19:26–32. doi: 10.1016/j.ymben.2013.05.003 CrossRefPubMedGoogle Scholar
  4. Bonnarme P, Gillet B, Sepulchre AM, Role C, Beloeil JC, Ducrocq C (1995) Itaconate biosynthesis in Aspergillus terreus. J Bacteriol 177(12):3573–3578PubMedPubMedCentralGoogle Scholar
  5. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254. doi: 10.1016/0003-2697(76)90527-3 CrossRefPubMedGoogle Scholar
  6. Cooper RA, Itiaba K, Kornberg HL (1965) The utilization of aconate and itaconate by Micrococcus sp. Biochem J 94:25–31. doi: 10.1042/bj0940025 CrossRefPubMedPubMedCentralGoogle Scholar
  7. Cooper RA, Kornberg HL (1964) The utilization of itaconate by Pseudomonas sp. Biochem J 91(1):82–91. doi: 10.1042/bj0910082 CrossRefPubMedPubMedCentralGoogle Scholar
  8. Dimroth P, Buckel W, Loyal R, Eggerer H (1977) Isolation and function of the subunits of citramalate lyase and formation of hybrids with the subunits of citrate lyase. Eur J Biochem 80(2):469–477. doi: 10.1111/j.1432-1033.1977.tb11902.x CrossRefPubMedGoogle Scholar
  9. Hevekerl A, Kuenz A, Vorlop KD (2014) Influence of the pH on the itaconic acid production with Aspergillus terreus. Appl Microbiol Biotechnol 98:10005–10012. doi: 10.1007/s00253-014-6047-2 CrossRefPubMedGoogle Scholar
  10. Hillier S, Charnetzky WT (1981) Glyoxylate bypass enzymes in Yersinia species and multiple forms of isocitrate lyase in Yersinia pestis. J Bacteriol 145(1):452–458PubMedPubMedCentralGoogle Scholar
  11. Honer Zu Bentrup K, Miczak A, Swenson DL, Russell DG (1999) Characterization of activity and expression of isocitrate lyase in Mycobacterium avium and Mycobacterium tuberculosis. J Bacteriol 181(23):7161–7167PubMedPubMedCentralGoogle Scholar
  12. Huang X, Lu X, Li JJ (2014a) Cloning, characterization and application of a glyceraldehyde-3-phosphate dehydrogenase promoter from Aspergillus terreus. J Ind Microbiol 41(3):585–592. doi: 10.1007/s10295-013-1385-0 CrossRefGoogle Scholar
  13. Huang X, Lu X, Li Y, Li X, Li J (2014b) Improving itaconic acid production through genetic engineering of an industrial Aspergillus terreus strain. Microb Cell Factories 13:119. doi: 10.1186/s12934-014-0119-y CrossRefGoogle Scholar
  14. Jaklitsch WM, Kubicek CP, Scrutton MC (1991) The subcellular organization of itaconate biosynthesis in Aspergillus terreus. J Gen Microbiol 137(3):533–539. doi: 10.1099/00221287-137-3-533 CrossRefGoogle Scholar
  15. Jakubowska J, Metodiewa D (1974) Studies on the metabolic pathway for itatartaric acid formation by Aspergillus terreus. II. Use of (−) citramalate, citraconate and itaconate by cell-free extracts. Acta Microbiol Pol B 6(2):51–61PubMedGoogle Scholar
  16. Li A, van Luijk N, ter Beek M, Caspers M, Punt P, van der Werf M (2011) A clone-based transcriptomics approach for the identification of genes relevant for itaconic acid production in Aspergillus. Fungal Genet Biol 48(6):602–611. doi: 10.1016/j.fgb.2011.01.013 CrossRefPubMedGoogle Scholar
  17. Lin YH, Li YF, Huang MC, Tsai YC (2004) Intracellular expression of Vitreoscilla hemoglobin in Aspergillus terreus to alleviate the effect of a short break in aeration during culture. Biotechnol Lett 26(13):1067–1072. doi: 10.1023/B:BILE.0000032964.15178.7c CrossRefPubMedGoogle Scholar
  18. Lubertozzi D, Keasling JD (2009) Developing Aspergillus as a host for heterologous expression. Biotechnol Adv 27(1):53–75. doi: 10.1016/j.biotechadv.2008.09.001 CrossRefPubMedGoogle Scholar
  19. Martin WR, Frigan F, Bergman EH (1961) Noninductive metabolism of itaconic acid by Pseudomonas and Salmonella species. J Bacteriol 82:905–908PubMedPubMedCentralGoogle Scholar
  20. Michelucci A, Cordes T, Ghelfi J, Pailot A, Reiling N, Goldmann O, Binz T, Wegner A, Tallam A, Rausell A, Buttini M, Linster CL, Medina E, Balling R, Hiller K (2013) Immune-responsive gene 1 protein links metabolism to immunity by catalyzing itaconic acid production. Proc Natl Acad Sci U S A 110(19):7820–7825. doi: 10.1073/pnas.1218599110 CrossRefPubMedPubMedCentralGoogle Scholar
  21. Okabe M, Lies D, Kanamasa S, Park EY (2009) Biotechnological production of itaconic acid and its biosynthesis in Aspergillus terreus. Appl Microbiol Biotechnol 84(4):597–606. doi: 10.1007/s00253-009-2132-3 CrossRefPubMedGoogle Scholar
  22. Reddy CS, Singh RP (2002) Enhanced production of itaconic acid from corn starch and market refuse fruits by genetically manipulated Aspergillus terreus SKR10. Bioresour Technol 85(1):69–71. doi: 10.1016/S0960-8524(02)00075-5 CrossRefPubMedGoogle Scholar
  23. Rychtera M, Wase DAJ (1981) The growth of Aspergillus terreus and the production of itaconic acid in batch and continuous cultures. The influence of pH. J Chem Technol Biotechnol 31(8):509–521. doi: 10.1002/jctb.503310168 CrossRefGoogle Scholar
  24. Sasikaran J, Ziemski M, Zadora PK, Fleig A, Berg IA (2014) Bacterial itaconate degradation promotes pathogenicity. Nat Chem Biol 10(5):371–377. doi: 10.1038/nchembio.1482 CrossRefPubMedGoogle Scholar
  25. Strelko CL, Lu W, Dufort FJ, Seyfried TN, Chiles TC, Rabinowitz JD, Roberts MF (2011) Itaconic acid is a mammalian metabolite induced during macrophage activation. J Am Chem Soc 133(41):16386–16389. doi: 10.1021/ja2070889 CrossRefPubMedPubMedCentralGoogle Scholar
  26. Tate H (1981) Assessing tumour markers. Br J Cancer 44(5):643–651Google Scholar
  27. Tevz G, Bencina M, Legisa M (2010) Enhancing itaconic acid production by Aspergillus terreus. Appl Microbiol Biotechnol 87(5):1657–1664. doi: 10.1007/s00253-010-2642-z CrossRefPubMedGoogle Scholar
  28. Wang SF, Adler J, Lardy HA (1961) The pathway of itaconate metabolism by liver mitochondria. J Biol Chem 236:26–30PubMedGoogle Scholar
  29. Willadsen P, Eggerer H (1975) Substrate stereochemistry of the enoyl-CoA hydratase reaction. Eur J Biochem 54(1):247–252. doi: 10.1111/j.1432-1033.1975.tb04134.x CrossRefPubMedGoogle Scholar
  30. Williams JO, Roche TE, McFadden BA (1971) Mechanism of action of isocitrate lyase from Pseudomonas indigofera. Biochemistry 10(8):1384–1390. doi: 10.1021/bi00784a017 CrossRefPubMedGoogle Scholar
  31. Yu CS, Chen YC, Lu CH, Hwang JK (2006) Prediction of protein subcellular localization. Proteins 64(3):643–651. doi: 10.1002/prot.21018 CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Mei Chen
    • 1
  • Xuenian Huang
    • 1
    • 2
  • Chengwei Zhong
    • 1
    • 2
  • Jianjun Li
    • 1
    • 3
  • Xuefeng Lu
    • 1
    Email author
  1. 1.Key Laboratory of Biofuels, Shandong Provincial Key Laboratory of Synthetic Biology, Qingdao Institute of Bioenergy and Bioprocess TechnologyChinese Academy of SciencesQingdaoChina
  2. 2.University of Chinese Academy of SciencesBeijingChina
  3. 3.National Key Laboratory of Biochemical Engineering, Institute of Process EngineeringChinese Academy of SciencesBeijingChina

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