The intracellular galactoglycome in Trichoderma reesei during growth on lactose
- 516 Downloads
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.
KeywordsTrichoderma reesei Lactose Galactoglycome Cellulase HPAEC-MS
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).
- 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
- 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
- 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
- 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
- Soper DS (2012) p-value calculator for correlation coefficients (online software). http://www.danielsoper.com/statcalc3