Advertisement

Applied Microbiology and Biotechnology

, Volume 97, Issue 12, pp 5447–5456 | Cite as

The intracellular galactoglycome in Trichoderma reesei during growth on lactose

  • Levente Karaffa
  • Leon Coulier
  • Erzsébet Fekete
  • Karin M. Overkamp
  • Irina S. Druzhinina
  • Marianna Mikus
  • Bernhard Seiboth
  • Levente Novák
  • Peter J. Punt
  • Christian P. Kubicek
Applied genetics and molecular biotechnology

Abstract

Lactose (1,4-0-β-d-galactopyranosyl-d-glucose) is used as a soluble carbon source for the production of cellulases and hemicellulases for—among other purposes—use in biofuel and biorefinery industries. The mechanism how lactose induces cellulase formation in T. reesei is enigmatic, however. Previous results from our laboratory raised the hypothesis that intermediates from the two galactose catabolic pathway may give rise to the accumulation of intracellular oligogalactosides that could act as inducer. Here we have therefore used high-performance anion-exchange chromatography–mass spectrometry to study the intracellular galactoglycome of T. reesei during growth on lactose, in T. reesei mutants impaired in galactose catabolism, and in strains with different cellulase productivities. Lactose, allo-lactose, and lactulose were detected in the highest amounts in all strains, and two trisaccharides (Gal-β-1,6-Gal-β-1,4-Glc/Fru and Gal-β-1,4-Gal-β-1,4-Glc/Fru) also accumulated to significant levels. Glucose and galactose, as well as four further oligosaccharides (Gal-β-1,3/1,4/1,6-Gal; Gal-β-1,2-Glc) were only detected in minor amounts. In addition, one unknown disaccharide (Hex-β-1,1-Hex) and four trisaccharides were also detected. The accumulation of the unknown hexose disaccharide was shown to correlate with cellulase formation in the improved mutant strains as well as the galactose pathway mutants, and Gal-β-1,4-Gal-β-1,4-Glc/Fru and two other unknown hexose trisaccharides correlated with cellulase production only in the pathway mutants, suggesting that these compounds could be involved in cellulase induction by lactose. The nature of these oligosaccharides, however, suggests their formation by transglycosylation rather than by glycosyltransferases. Based on our results, the obligate nature of both galactose catabolic pathways for this induction must have another biochemical basis than providing substrates for inducer formation.

Keywords

Trichoderma reesei Lactose Galactoglycome Cellulase HPAEC-MS 

Notes

Acknowledgments

This work was supported by grants from the Austrian Science Foundation (FWF P-19143 and FWF P24219) to CPK and BS, respectively. Research at the University of Debrecen has been supported by the OTKA (Hungarian Scientific Research Fund; grants K67667 and K1006600) and by the TÁMOP-4.2.2/B-10/-1-2010-0024 Project. EF is a recipient of a Bólyai János Research Scholarship (BO/00519/09/8).

References

  1. Bruggink C, Maurer R, Herrmann H, Cavalli S, Hoefler FJ (2005) Analysis of carbohydrates by anion exchange chromatography and mass spectrometry. J Chromatography A 1085:104–109CrossRefGoogle Scholar
  2. Bucior I, Burger MM (2004) Carbohydrate–carbohydrate interaction as a major force initiating cell–cell recognition. Glycoconj J 21:111–123CrossRefGoogle Scholar
  3. Coulier L, Timmermans J, Richard B, Van Den Dool R, Haaksman I, Klarenbeek B, Slaghek T, Van Dongen WJ (2009) In-depth characterization of prebiotic galacto-oligosaccharides by a combination of analytical techniques. J Agr Food Chem 57:8488–8495CrossRefGoogle Scholar
  4. Eveleigh DE, Montenecourt BS (1979) Increasing yields of extracellular enzymes. Adv Appl Microbiol 25:57–74CrossRefGoogle Scholar
  5. Fekete E, Seiboth B, Kubicek CP, Szentirmai A, Karaffa L (2008) Lack of aldose 1-epimerase in Hypocrea jecorina (anamorph Trichoderma reesei): a key to cellulase gene expression on lactose. Proc Natl Acad Sci USA 105:7141–7146CrossRefGoogle Scholar
  6. Hartl L, Kubicek CP, Seiboth B (2007) Induction of the gal pathway and cellulase genes involves no transcriptional inducer function of the galactokinase in Hypocrea jecorina. J Biol Chem 282:18654–18659CrossRefGoogle Scholar
  7. Jörgensen RG, Mäder P, Fließbach A (2010) Long-term effects of organic farming on fungal and bacterial residues in relation to microbial energy metabolism. Biol Fert Soils 46:303–307CrossRefGoogle Scholar
  8. Karaffa L, Fekete E, Gamauf C, Szentirmai A, Kubicek CP, Seiboth B (2006) D-Galactose induces cellulase gene expression in Hypocrea jecorina at low growth rates. Microbiology 152:1507–1514CrossRefGoogle Scholar
  9. Lairson LL, Henrissat B, Davies GJ, Withers SG (2008) Glycosyltransferases: structures, functions, and mechanisms. Annu Rev Biochem 77:521–55CrossRefGoogle Scholar
  10. Lamsal BP (2012) Production, health aspects and potential food uses of dairy prebiotic galactooligosaccharides. J Sci Food Agr 92:2020–2028CrossRefGoogle Scholar
  11. Le Crom S, Schackwitz W, Penacchio L, Magnuson J, Culley D, Collet J, Martin J, Druzhinina IS, Mathis H, Monot F, Seiboth B, Cherry B, Rey M, Berka R, Kubicek CP, Baker SE, Margeot A (2009) Tracking the roots of cellulase hyperproduction by the fungus Trichoderma reesei using massively parallel DNA sequencing. Proc Natl Acad Sci USA 106:16151–16156CrossRefGoogle Scholar
  12. Lu L, Xu S, Jin L, Zhang D, Li Y, Xiao M (2012) Synthesis of galactosyl sucralose by β-galactosidase from Lactobacillus bulgaricus L3. Food Chem 134:269–275CrossRefGoogle Scholar
  13. Martinez D, Berka RM, Henrissat B, Saloheimo M, Arvas M, Baker SE, Chapman J, Chertkov O, Coutinho PM, Cullen D, Danchin EG, Grigoriev IV, Harris P, Jackson M, Kubicek CP, Han CS, Ho I, Larrondo LF, de Leon AL, Magnuson JK, Merino S, Misra M, Nelson B, Putnam N, Robbertse B, Salamov AA, Schmoll M, Terry A, Thayer N, Westerholm-Parvinen A, Schoch CL, Yao J, Barabote R, Nelson MA, Detter C, Bruce D, Kuske CR, Xie G, Richardson P, Rokhsar DS, Lucas SM, Rubin EM, Dunn-Coleman N, Ward M, Brettin TS (2008) Genome sequencing and analysis of the biomass-degrading fungus Trichoderma reesei (syn. Hypocrea jecorina). Nat Biotechnol 26:553–560CrossRefGoogle Scholar
  14. Menon V, Rao M (2012) Trends in bioconversion of lignocellulose: biofuels, platform chemicals & biorefinery concept. Prog Energy Comb Sci 38:522–550CrossRefGoogle Scholar
  15. Nagai A, Yamamoto T, Wariishi H (2012) Identification of fructo- and malto-oligosaccharides in cured tobacco leaves (Nicotiana tabacum). J Agri Food Chem 60:6606–6612CrossRefGoogle Scholar
  16. Okatch H, Torto N, Armateifio J (2003) Characterisation of legumes by enzymatic hydrolysis, microdialysis sampling, and micro-high-performance anion-exchange chromatography with electrospray ionization mass spectrometry. J Chromatography A 992:67–74CrossRefGoogle Scholar
  17. Prasad S, Roy I (2010) Effect of disaccharides on the stabilization of bovine trypsin against detergent and autolysis. Biotechnol Prog 26:627–635CrossRefGoogle Scholar
  18. Richardson S, Cohen A, Gorton L (2001) High-performance chromatography-electrospray mass spectrometry for investigation of the substituent distribution in hydroxypropylated potato amylopectin starch. J Chromatography A 917:111–121CrossRefGoogle Scholar
  19. Rodriguez-Fernandez M, Cardelle-Cobas A, Villamiel M, Banga JR (2011) Detailed kinetic model describing new oligosaccharides synthesis using different β-galactosidases. J Biotechnol 153:116–124CrossRefGoogle Scholar
  20. Roelfsema WA, Kuster BFM, Heslinga MC, Pluim H, Verhage M (2010) Lactose and derivatives. Ullmann’s encyclopedia of industrial chemistry, 7th edn. Wiley, New YorkGoogle Scholar
  21. Rodgers JL, Nicewander WA (1988) Thirteen ways to look at the correlation coefficient. American Stat 42:59–66CrossRefGoogle Scholar
  22. Röhr M, Zehentgruber O, Kubicek CP (1981) Kinetics of biomass formation and citric acid production by Aspergillus niger on a pilot plant scale. Biotechnol Bioeng 23:2433–2445CrossRefGoogle Scholar
  23. Ruijter GJG, Visser J (1996) Determination of intermediary metabolites in Aspergillus niger. J Microbiol Methods 25:295–302CrossRefGoogle Scholar
  24. Rumbold K, Okatch H, Torto N, Siika-Aho M, Gubitz G, Robra K-H, Prior B (2002) Monitoring on-line desalted lignocellulosic hydrolysates by microdialysis sampling micro-high performance anion exchange chromatography with integrated pulsed electrochemical detection/mass spectrometry. Biotechnol Bioeng 78:822–828CrossRefGoogle Scholar
  25. Sangwan V, Tomar SK, Singh RRB, Singh AK, Ali B (2011) Galactooligosaccharides: novel components of designer foods. J Food Sci 76:R103–R111CrossRefGoogle Scholar
  26. Seiboth B, Hartl L, Pail M, Fekete E, Karaffa L, Kubicek CP (2004) The galactokinase of Hypocrea jecorina is essential for cellulase induction by lactose but dispensable for growth on D-galactose. Mol Microbiol 51:1015–1025CrossRefGoogle Scholar
  27. Seiboth B, Gamauf C, Pail M, Hartl L, Kubicek CP (2007) The D-xylose reductase of Hypocrea jecorina is the major aldose reductase in pentose and D-galactose catabolism and necessary for β-galactosidase and cellulase induction by lactose. Mol Microbiol 66:890–900CrossRefGoogle Scholar
  28. Soper DS (2012) p-value calculator for correlation coefficients (online software). http://www.danielsoper.com/statcalc3

Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  • Levente Karaffa
    • 1
  • Leon Coulier
    • 2
  • Erzsébet Fekete
    • 1
  • Karin M. Overkamp
    • 2
  • Irina S. Druzhinina
    • 3
    • 4
  • Marianna Mikus
    • 3
    • 4
  • Bernhard Seiboth
    • 3
    • 4
  • Levente Novák
    • 5
  • Peter J. Punt
    • 2
  • Christian P. Kubicek
    • 3
    • 4
  1. 1.Department of Biochemical Engineering, Faculty of Science and TechnologyUniversity of DebrecenDebrecenHungary
  2. 2.TNOZeistthe Netherlands
  3. 3.Research Area Biotechnology and Microbiology, Institute of Chemical EngineeringTU WienViennaAustria
  4. 4.Austrian Center of Industrial Biotechnology (ACIB), c/o TU WienViennaAustria
  5. 5.Department of Colloid and Environmental Chemistry, Faculty of Science and TechnologyUniversity of DebrecenDebrecenHungary

Personalised recommendations