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Enhanced Production of Itaconic Acid through Development of Transformed Fungal Strains of Aspergillus terreus

  • Shin, Woo-Shik (Green Materials and Process R&BD Group, Korea Institute of Industrial Technology) ;
  • Park, Boonyoung (Green Materials and Process R&BD Group, Korea Institute of Industrial Technology) ;
  • Lee, Dohoon (Green Materials and Process R&BD Group, Korea Institute of Industrial Technology) ;
  • Oh, Min-Kyu (Department of Chemical and Biological Engineering, Korea University) ;
  • Chun, Gie-Taek (College of Biomedical Science, Kangwon National University) ;
  • Kim, Sangyong (Green Materials and Process R&BD Group, Korea Institute of Industrial Technology)
  • Received : 2016.11.18
  • Accepted : 2016.12.06
  • Published : 2017.02.28

Abstract

Metabolic engineering with a high-yielding mutant, A. terreus AN37, was performed to enhance the production of itaconic acid (IA). Reportedly, the gene cluster for IA biosynthesis is composed of four genes: reg (regulator), mtt (mitochondrial transporter), cad (cis-aconitate decarboxylase), and mfs (membrane transporter). By overexpressing each gene of the IA gene cluster in A. terreus AN37 transformed by the restriction enzyme-mediated integration method, several transformants showing high productivity of IA were successfully obtained. One of the AN37/cad transformants could produce a very high amount of IA (75 g/l) in shake-flask cultivations, showing an average of 5% higher IA titer compared with the high-yielding control strain. Notably, in the case of the mfs transformants, a maximal increase of 18.3% in IA production was observed relative to the control strain under the identical fermentation conditions. Meanwhile, the overexpression of reg and mtt genes showed no significant improvements in IA production. In summary, the overexpressed cis-aconitate decarboxylase (CAD) and putative membrane transporter (MFS) appeared to have positive influences on the enhanced IA productivity of the respective transformant. The maximal increases of 13.6~18.3% in IA productivity of the transformed strains should be noted, since the parallel mother strain used in this study is indeed a very high-performance mutant that has been obtained through intensive rational screening programs in our laboratory.

Keywords

References

  1. Tevz G, Bencina M, Legisa M. 2010. Enhancing itaconic acid production by Aspergillus terreus. Appl. Microbiol. Biotechnol. 87: 1657-1664. https://doi.org/10.1007/s00253-010-2642-z
  2. Steiger M, Blumhoff M, Mattanovich D, Sauer M. 2013. Biochemistry of microbial itaconic acid production. Front. Microbiol. 4: 23.
  3. Pedroso GB, Montipo S, Mario DAN, Alves SH, Martins AF. 2016. Building block itaconic acid from left-over biomass. Biomass Convers. Biorefinery DOI: 10.1007/s1339901602101.
  4. Clomburg JM, Gonzalez R. 2010. Biofuel production in Escherichia coli: the role of metabolic engineering and synthetic biology. Appl. Microbiol. Biotechnol. 86: 419-434. https://doi.org/10.1007/s00253-010-2446-1
  5. Tong X, Ma Y, Li Y. 2010. Biomass into chemicals: conversion of sugars to furan derivatives by catalytic processes. Appl. Catal. A Gen. 385: 1-13. https://doi.org/10.1016/j.apcata.2010.06.049
  6. Straat L, Vernooij M, Lammers M, Berg W, Schonewille T, Cordewener J, et al. 2014. Expression of the Aspergillus terreus itaconic acid biosynthesis cluster in Aspergillus niger. Microb. Cell Fact. 13: 11. https://doi.org/10.1186/1475-2859-13-11
  7. Bonnarme P, Gillet B, Sepulchre A, Role C, Beloeil J, Ducrocq C. 1995. Itaconate biosynthesis in Aspergillus terreus. J. Bacteriol. 177: 3573-3578. https://doi.org/10.1128/jb.177.12.3573-3578.1995
  8. Klement T, Buchs J. 2013. Itaconic acid - a biotechnological process in change. Bioresour. Technol. 135: 422-431. https://doi.org/10.1016/j.biortech.2012.11.141
  9. Lee JW, Kim HU, Choi S, Yi J, Lee SY. 2011. Microbial production of building block chemicals and polymers. Curr. Opin. Biotechnol. 22: 758-767. https://doi.org/10.1016/j.copbio.2011.02.011
  10. Werpy T, Petersen G. 2004. Top Value Added Chemicals from Biomass - Volume I: Results of Screening for Potential Candidates from Sugars and Synthesis Gas. Department of Energy, Oak Ridge, TN, USA.
  11. Okabe M, Lies D, Kanamasa S, Park EY. 2009. Biotechnological production of itaconic acid and its biosynthesis in Aspergillus terreus. Appl. Microbiol. Biotechnol. 84: 597-606. https://doi.org/10.1007/s00253-009-2132-3
  12. Li A, Luijk N, Beek M, Caspers M, Punt P, Werf M. 2011. A clone-based transcriptomics approach for the identification of genes relevant for itaconic acid production in Aspergillus. Fungal Genet. Biol. 48: 602-611. https://doi.org/10.1016/j.fgb.2011.01.013
  13. Jaklitsch WM, Kubicek CP, Scrutton MC. 1991. The subcellular organization of itaconate biosynthesis in Aspergillus terreus. J. Gen. Microbiol. 137: 533-539. https://doi.org/10.1099/00221287-137-3-533
  14. Li A, Caspers M, Punt P. 2013. A systems biology approach for the identification of target genes for the improvement of itaconic acid production in Aspergillus species. BMC Res. Notes 6: 1-5. https://doi.org/10.1186/1756-0500-6-1
  15. Li A, Pfelzer N, Zuijderwijk R, Punt P. 2012. Enhanced itaconic acid production in Aspergillus niger using genetic modification and medium optimization. BMC Biotechnol. 12: 57. https://doi.org/10.1186/1472-6750-12-57
  16. Holz M, Forster A, Mauersberger S, Barth G. 2009. Aconitase overexpression changes the product ratio of citric acid production by Yarrowia lipolytica. Appl. Microbiol. Biotechnol. 81: 1087-1096. https://doi.org/10.1007/s00253-008-1725-6
  17. Kanamasa S, Dwiarti L, Okabe M, Park E. 2008. Cloning and functional characterization of the cis-aconitic acid decarboxylase (CAD) gene from Aspergillus terreus. Appl. Microbiol. Biotechnol. 80: 223-229. https://doi.org/10.1007/s00253-008-1523-1
  18. Shin WS, Lee D, Kim S, Jeong YS, Chun GT. 2013. Application of scale-up criterion of constant oxygen mass transfer coefficient (kLa) for production of itaconic acid in a 50 L pilot-scale fermentor by fungal cells of Aspergillus terreus. J. Microbiol. Biotechnol. 23: 1445-1453. https://doi.org/10.4014/jmb.1307.07084
  19. Park SM. 2013. Development of high-yielding strains and statistical medium optimization for industial production of itaconic acid. Kangwon National University, Korea.
  20. Shin WS. 2011. Development of high yielding strain and establishment of bioprocess system or mass production of itaconic acid by Aspergillus terreus cells. PhD thesis. Kangwon National University, Korea.
  21. Sambrook J, Fritsch EF, Maniatis T. 1989. Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.
  22. Thon MR, Nuckles EM, Vaillancourt LJ. 2000. Restriction enzyme-mediated integration used to produce pathogenicity mutants of Colletotrichum graminicola. Mol. Plant Microbe Interact. 13: 1356-1365. https://doi.org/10.1094/MPMI.2000.13.12.1356
  23. Schiestl RH, Petes TD. 1991. Integration of DNA fragments by illegitimate recombination in Saccharomyces cerevisiae. Proc. Natl. Acad. Sci. USA 88: 7585-7589. https://doi.org/10.1073/pnas.88.17.7585
  24. Brown JS, Aufauvre-Brown A, Holden DW. 1998. Insertional mutagenesis of Aspergillus fumigatus. Mol. Gen. Genet. 259: 327-335. https://doi.org/10.1007/s004380050819
  25. Bolker M, Bohnert HU, Braun KH, Gorl J, Kahmann R. 1995. Tagging pathogenicity genes in Ustilago maydis by restriction enzyme-mediated integration (REMI). Mol. Gen. Genet. 248: 547-552. https://doi.org/10.1007/BF02423450
  26. Huang X, Lu X, Li Y, Li X, Li J-J. 2014. Improving itaconic acid production through genetic engineering of an industrial Aspergillus terreus strain. Microb. Cell Fact. 13: 1-9. https://doi.org/10.1186/1475-2859-13-1
  27. Gennuso F, Fernetti C, Tirolo C, Testa N, L'Episcopo F, Caniglia S, et al. 2004. Bilirubin protects astrocytes from its own toxicity by inducing up-regulation and translocation of multidrug resistance-associated protein 1 (Mrp1). Proc. Natl. Acad. Sci. USA 101: 2470-2475. https://doi.org/10.1073/pnas.0308452100
  28. Hipfner DR, Gauldie SD, Deeley RG, Cole SP. 1994. Detection of the M(r) 190,000 multidrug resistance protein, MRP, with monoclonal antibodies. Cancer Res. 54: 5788-5792.
  29. Rajagopal A, Pant AC, Simon SM, Chen Y. 2002. In vivo analysis of human multidrug resistance protein 1 (MRP1) activity using transient expression of fluorescently tagged MRP1. Cancer Res. 62: 391-396.
  30. Koley D, Bard AJ. 2010. Triton X-100 concentration effects on membrane permeability of a single HeLa cell by scanning electrochemical microscopy (SECM). Proc. Natl. Acad. Sci. USA 107: 16783-16787. https://doi.org/10.1073/pnas.1011614107
  31. Burg RW, Stapley EO. 1989. Isolation and characterization of the producing organism, pp. 24-32. In Campbell WC (ed.). Ivermectin and Abamectin. Springer New York, New York, NY.
  32. Studer MH, DeMartini JD, Brethauer S, McKenzie HL, Wyman CE. 2010. Engineering of a high-throughput screening system to identify cellulosic biomass, pretreatments, and enzyme formulations that enhance sugar release. Biotechnol. Bioeng. 105: 231-238. https://doi.org/10.1002/bit.22527

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