Advertisement

Purification and characterization of an endo-xylanase from Trichoderma sp., with xylobiose as the main product from xylan hydrolysis

  • Li-Hao Fu
  • Nan Jiang
  • Cheng-Xi Li
  • Xue-Mei Luo
  • Shuai ZhaoEmail author
  • Jia-Xun FengEmail author
Original Paper

Abstract

Fungal endo-β-1,4-xylanases (endo-xylanases) can hydrolyze xylan into xylooligosaccharides (XOS), and have potential biotechnological applications for the exploitation of natural renewable polysaccharides. In the current study, we aimed to screen and characterize an efficient fungal endo-xylanase from 100 natural humus-rich soil samples collected in Guizhou Province, China, using extracted sugarcane bagasse xylan (SBX) as the sole carbon source. Initially, 182 fungal isolates producing xylanases were selected, among which Trichoderma sp. strain TP3-36 was identified as showing the highest xylanase activity of 295 U/mL with xylobiose (X2) as the main product when beechwood xylan was used as substrate. Subsequently, a glycoside hydrolase family 11 endo-xylanase, TXyn11A, was purified from strain TP3-36, and its optimal pH and temperature for activity against beechwood xylan were identified to be 5.0 and 55 °C, respectively. TXyn11A was stable across a broad pH range (3.0–10.0), and exhibited strict substrate specificity, including xylan from beechwood, wheat, rye, and sugarcane bagasse, with Km and Vmax values of 5 mg/mL and 1250 μmol/mg min, respectively, toward beechwood xylan. Intriguingly, the main product obtained from hydrolysis of beechwood xylan by TXyn11A was xylobiose, whereas SBX hydrolysis resulted in both X2 and xylotriose. Overall, these characteristics of the endo-xylanase TXyn11A indicate several potential industrial applications.

Keywords

Endo-xylanase Xylobiose Sugarcane bagasse xylanase Trichoderma sp. 

Abbreviations

CMC

Carboxymethylcellulose

DNS

3,5-Dinitrosalicylic acid

DTT

Dithiothreitol

GH

Glycoside hydrolase

HPLC

High-performance liquid chromatography

ITS

Internal transcribed spacer

PDA

Potato dextrose agar

SB

Sugarcane bagasse

SBX

SB xylan

SMART

Simple modular architecture research tool

Tm

Temperature

XOS

Xylooligosaccharides

X1

Xylose

X2

Xylobiose

X3

Xylotriose

X4

Xylotetraose

Notes

Acknowledgements

We would like to thank Mu-Qing Zhang group from State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources, Guangxi University, China, for providing raw SB materials. This work was financially supported by Grants from the Autonomous Research Project of State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources (SKLCUSA-a201902, SKLCUSA-a201923), the Guangxi BaGui Scholars Program Foundation (Grant No. 2011A001), the Guangxi Natural Science Foundation (Grant No. 2012GXNSFGA060005), Training Program for 1000 Young and Middle-Aged Key Teachers in Guangxi at 2019, and the ‘One Hundred Person’ Project of Guangxi.

Supplementary material

11274_2019_2747_MOESM1_ESM.pdf (1.5 mb)
Supplementary file1 (PDF 1563 kb)

References

  1. Aachary AA, Prapulla SG (2011) Xylooligosaccharides (XOS) as an emerging prebiotic: microbial synthesis, utilization, structural characterization, bioactive properties, and applications. Compr Rev Food Sci Food Saf 10:2–16Google Scholar
  2. Bhatia L, Sharma A, Bachheti RK, Chandel AK (2019) Lignocellulose derived functional oligosaccharides: production, properties, and health benefits. Prep Biochem Biotechnol.  https://doi.org/10.1080/10826068.2019.1608446 CrossRefPubMedGoogle Scholar
  3. Bhardwaj N, Kumar B, Agarwal K, Chaturvedi V, Verma P (2019) Purification and characterization of a thermo-acid/alkali stable xylanases from Aspergillus oryzae LC1 and its application in Xylo-oligosaccharides production from lignocellulosic agricultural wastes. Int J Biol Macromol 122:1191–1202PubMedGoogle Scholar
  4. Bian J, Peng F, Peng XP, Peng P, Xu F, Sun RC (2013) Structural features and antioxidant activity of xylooligosaccharides enzymatically produced from sugarcane bagasse. Bioresour Technol 127:236–241PubMedGoogle Scholar
  5. Biely P, Puchart V, Stringer MA, Krogh KBRM (2014) Trichoderma reesei XYN VI—a novel appendage dependent eukaryotic glucuronoxylan hydrolases. FEBS J 281:3894–3903Google Scholar
  6. Biely P, Singh S, Puchart V (2016) Towards enzymatic breakdown of complex plant xylan structures: state of the art. Biotechnol Adv 34:1260–1274Google Scholar
  7. Brienzo M, Carvalho W, Milagres AMF (2010) Xylooligosaccharides production from alkali-pretreated sugarcane bagasse using xylanases from Thermoascus aurantiacus. Appl Biochem Biotechnol 162:1195–1205PubMedGoogle Scholar
  8. Chaverri P (2015) Systematics of the Trichoderma harzianum species complex and the re-identification of commercial biocontrol strains. Mycologia 107:558–590PubMedPubMedCentralGoogle Scholar
  9. Chong SL, Virkki L, Maaheimo H, Juvonen M, Derba-Maceluch M, Koutaniemi S, Roach M, Sundberg B, Tuomainen P, Mellerowicz EJ, Tenkanen M (2014) O-acetylation of glucuronoxylan in Arabidopsis thaliana wild type and its change in xylan biosynthesis mutants. Glycobiology 24:494–506PubMedGoogle Scholar
  10. Chung YC, Hsieh CP, Chan YC (2002) Effect of xylooligosaccharides on the intestinal properties of ICR mice. Taiwan J Agric Chem Food Sci 40:377–384Google Scholar
  11. Chung YC, Hsu CK, Ko CY, Chan YC (2007) Dietary intake of xylooligosaccharides improves the intestinal microbiota, fecal moisture, and pH value in the elderly. Nutr Res 27:756–761Google Scholar
  12. Ding C, Li M, Hu Y (2018) High-activity production of xylanase by Pichia stipitis: purification, characterization, kinetic evaluation and xylooligosaccharides production. Int J Biol Macromol 117:72–77PubMedGoogle Scholar
  13. Ebringerova A, Heinze T (2000) Xylan and xylan derivatives-biopolymers with valuable properties, 1. Naturally occurring xylans structures, isolation procedures and properties. Macromol Rapid Commun 21:542–556Google Scholar
  14. Ebringerová A, Hromádková Z, Heinze T (2005) Hemicellulose. In: Heinze T (ed) Polysaccharides I: structure, characterization and use. Springer, Berlin, pp 1–67Google Scholar
  15. Espinoza K, Eyzaguirre J (2018) Identification, heterologous expression and characterization of a novel glycoside hydrolase family 30 xylanase from the fungus Penicillium purpurogenum. Carbohydr Res 468:45–50PubMedGoogle Scholar
  16. Evtuguin DV, Tomás JL, Silva AMS, Neto CP (2003) Characterization of an acetylated heteroxylan from Eucalyptus globulus Labill. Carbohydr Res 338:597–604PubMedGoogle Scholar
  17. Fang HY, Chang SM, Lan CH, Fang TJ (2008) Purification and characterization of a xylanase from Aspergillus carneus M34 and its potential use in photoprotectant preparation. Process Biochem 43:49–55Google Scholar
  18. Fernández-Espinar M, Piñaga F, de Graaff L, Visser J, Ramón D, Vallés S (1994) Purification, characterization and regulation of the synthesis of an Aspergillus nidulans acidic xylanase. Appl Microbiol Biotechnol 42:555–562Google Scholar
  19. Grigoriev IV, Nikitin R, Haridas S, Kuo A, Ohm R, Otillar R, Riley R, Salamov A, Zhao X, Korzeniewski F, Smirnova T, Nordberg H, Dubchak I, Shabalov I (2014) MycoCosm portal: gearing up for 1000 fungal genomes. Nucleic Acids Res 42(1):D699–D704PubMedGoogle Scholar
  20. Jiang SX, Zhao S, Lu CY, Xue JL, Duan CJ, Feng JX (2017) A combined process is used for efficient isolation and purification of xylobiose from xylanase-hydrolysed sugarcane bagasse xylan hydrolysate. Ind Crop Prod 109:637–643Google Scholar
  21. Jing L, Zhao S, Xue JL, Zhang Z, Yang Q, Xian L, Feng JX (2015) Isolation and characterization of a novel Penicillium oxalicum strain Z1–3 with enhanced cellobiohydrolase production using cellulase-hydrolyzed sugarcane bagasse as carbon source. Ind Crop Prod 77:666–675Google Scholar
  22. Juturu V, Wu JC (2012) Microbial xylanases: engineering, production and industrial application. Biotechnol Adv 30:1219–1227Google Scholar
  23. Kajihara M, Kato S, Konishi M, Yamagishi Y, Horie Y, Ishii H (2000) Xylooligosaccharide decreases blood ammonia levels in patients with liver cirrhosis. Am J Gastroenterol 95:2514Google Scholar
  24. Karlsson EN, Schmitz E, Linares-Pastén JA, Adlercreutz P (2018) Endo-xylanases as tools for production of substituted xylooligosaccharides with prebiotic properties. Appl Microbiol Biotechnol 102:9081–9088Google Scholar
  25. Katsimpouras C, Dedes G, Thomaidis NS, Topakas E (2019) A novel fungal GH30 xylanase with xylobiohydrolase auxiliary activity. Biotechnol Biofuels 12:120PubMedPubMedCentralGoogle Scholar
  26. Kumar S, Stecher G, Tamura K (2016) MEGA7: molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol Biol Evol 33:1870–1874PubMedGoogle Scholar
  27. Kumar V, Dangi AK, Shukla P (2018) Engineering thermostable microbial xylanases toward its industrial applications. Mol Biotechnol 60:226–235PubMedGoogle Scholar
  28. Li M, Yang Q (2007) Isolation and characterization of a β-tubulin gene from Trichoderma harzianum. Biochem Genet 45:529–534PubMedGoogle Scholar
  29. Li Q, Wu QH, Sun BG, Yang R, Hou X, Teng C, Zhang CN, Li XT (2018) Effect of disulfide bridge on hydrolytic characteristics of xylanase from Penicillium janthinellum. Int J Biol Macromol 120:405–413PubMedGoogle Scholar
  30. Linares-Pasten JA, Aronsson A, Karlsson EN (2018) Structural considerations on the use of endo-xylanases for the production of prebiotic xylooligosaccharides from biomass. Curr Protein Pept Sci 19:48–67PubMedPubMedCentralGoogle Scholar
  31. Liu TJ, Williams DL, Pattathil S, Li MY, Hahn MG, Hodge DB (2014) Coupling alkaline pre-extraction with alkaline-oxidative post-treatment of corn stover to enhance enzymatic hydrolysis and fermentability. Biotechnol Biofuels 7:48PubMedPubMedCentralGoogle Scholar
  32. Luo HY, Yang J, Li J, Shi PJ, Huang HQ, Bai YG, Fan YL, Yao B (2010) Molecular cloning and characterization of the novel acidic xylanase XYLD from Bispora sp. MEY-1 that is homologous to family 30 glycosyl hydrolases. Appl Microbiol Biotechnol 86:1829–1839PubMedGoogle Scholar
  33. Mano MCR, Neri-Numa IA, da Silva JB, Paulino BN, Pessoa MG, Pastore GM (2018) Oligosaccharide biotechnology: an approach of prebiotic revolution on the industry. Appl Microbiol Biotechnol 102:17–37PubMedGoogle Scholar
  34. Mazumder K, Peña MJ, O’Neill MA, York WS (2012) Structural characterization of the heteroxylans from poplar and switchgrass. Methods Mol Biol 908:215–228PubMedGoogle Scholar
  35. Miller GL (1959) Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal Chem 31:426–428Google Scholar
  36. Moura P, Barata R, Carvalheiro F, Girio F, Loureiroias MC, Esteves MP (2007) In vitro fermentation of xylo-oligosaccharides from corn cobs autohydrolysis by Bifidobacterium and Lactobacillus strains. Lebensm Wiss Technol 40:963–972Google Scholar
  37. Naidu DS, Hlangothi SP, John MJ (2018) Bio-based products from xylan: a review. Carbohydr Polym 179:28–41PubMedGoogle Scholar
  38. Nakamichi Y, Fouquet T, Ito S, Matsushika A, Inoue H (2019a) Mode of action of GH30-7 reducing-end xylose-releasing exoxylanase A (Xyn30A) from the filamentous fungus Talaromyces cellulolyticus. Appl Environ Microbiol 85:e00552–e619PubMedGoogle Scholar
  39. Nakamichi Y, Fouquet T, Ito S, Watanabe M, Matsushika A, Inoue H (2019b) Structural and functional characterization of a bifunctional GH30-7 xylanase B from the filamentous fungus Talaromyces cellulolyticus. J Biol Chem 294:4065–4078PubMedGoogle Scholar
  40. OECD (2017) Safety considerations for biotechnology 1992. Organization for Economic Co-operation and Development. https://www.oecd.org/sti/biotech/2375496.pdf. Accessed 28 July 2017
  41. Paes G, Berrin JG, Beaugrand J (2012) GH11 xylanases: structure/function/properties relationships and applications. Biotechnol Adv 30:564–592PubMedGoogle Scholar
  42. Peña MJ, Zhong RQ, Zhou GK, Richardson EA, O’Neill MA, Darvill AG, York WS, Ye ZH (2007) Arabidopsis irregular xylem8 and irregular xylem9: implications for the complexity of glucuronoxylan biosynthesis. Plant Cell 19:549–563PubMedGoogle Scholar
  43. Polizeli MLTM, Rizzatti ACS, Monti R, Terenzi HF, Jorge JA, Amorim DS (2005) Xylanases from fungi: properties and industrial applications. Appl Microbiol Biotechnol 67:577–591PubMedGoogle Scholar
  44. Romanowska I, Polak J, Bielecki S (2006) Isolation and properties of Aspergillus niger IBT-90 xylanase for bakery. Appl Microbiol Biotechnol 69:665–671PubMedGoogle Scholar
  45. Seifert E (2014) Origin Pro 9.1: scientific data analysis and graphing software—software review. J Chem Inf Model 54:1552PubMedGoogle Scholar
  46. Silva LAO, Terrasan CRF, Carmona EC (2015) Purification and characterization of xylanases from Trichoderma inhamatum. Electron J Biotechnol 18:307–313Google Scholar
  47. Smith PJ, Wang HT, York WS, Peña MJ, Urbanowicz BR (2017) Designer biomass for next-generation biorefineries: leveraging recent insights into xylan structure and biosynthesis. Biotechnol Biofuels 10:286PubMedPubMedCentralGoogle Scholar
  48. Sporck D, Reinoso FAM, Rencoret J, Gutiérrez A, Del Rio JC, Ferrza A, Milagres AMF (2017) Xylan extraction from pretreated sugarcane bagasse using alkaline and enzymatic approaches. Biotechnol Biofuels 10:296PubMedPubMedCentralGoogle Scholar
  49. Tan LUL, Wong KKY, Saddler JN (1985) Functional characteristics of two d-xylanases purified from Trichoderma harzianum. Enzyme Microb Technol 7:431–436Google Scholar
  50. Tan SS, Li DY, Jiang ZQ, Zhu YP, Shi B, Li LT (2008) Production of xylobiose from the autohydrolysis explosion liquor of corncob using Thermotoga maritima xylanase B (XynB) immobilized on nickel-chelated Eupergit C. Bioresour Technol 99:200–204PubMedGoogle Scholar
  51. Teng C, Yan Q, Jiang Z, Fan G, Shi B (2010) Production of xylooligosaccharides from the steam explosion liquor of corncobs coupled with enzymatic hydrolysis using a thermostable xylanase. Bioresour Technol 101:7679–7682PubMedGoogle Scholar
  52. Tenkanen M, Vršanská M, Siika-Aho M, Wong DW, Puchart V, Penttilä M, Saloheimo M, Biely P (2013) Xylanase XYN IV from Trichoderma reesei showing exo- and endo-xylanase activity. FEBS J 280:285–301PubMedGoogle Scholar
  53. Tischler BY, Hohl TM (2019) Menacing mold: recent advances in Aspergillus pathogenesis and host defense. J Mol Biol.  https://doi.org/10.1016/j.jmb.2019.03.027 CrossRefPubMedGoogle Scholar
  54. Uday USP, Choudhury P, Bandyopadhyay TK, Bhunia B (2015) Classification, mode of action and production strategy of xylanase and its application for biofuel production from water hyacinth. Int J Biol Macromol 82:1041–1054PubMedGoogle Scholar
  55. White TJ, Bruns T, Lee S, Taylor J (1990) Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innis MA, Gelfand DH, Shinsky JJ, White TJ (eds) PCR protocols: a guide to methods and applications. Academic, San Diego, pp 315–322Google Scholar
  56. Wong KKY, Tan LUL, Saddler JN (1986) Purification of a third distinct xylanase from the xylanolytic system of Trichoderma harzianum. Can J Microbiol 32:570–576Google Scholar
  57. Xian L, Feng JX (2018) Purification and biochemical characterization of a novel mesophilic glucoamylase from Aspergillus tritici. Int J Biol Macromol 107:1122–1130PubMedGoogle Scholar
  58. Xian L, Wang F, Luo X, Feng YL, Feng JX (2015) Purification and characterization of a highly efficient calcium-independent α-amylase from Talaromyces pinophilus 1-95. PLoS ONE 10:e0121531PubMedPubMedCentralGoogle Scholar
  59. Xu QS, Yan YS, Feng JX (2016) Efficient hydrolysis of raw starch and ethanol fermentation: a novel raw starch-digesting glucoamylase from Penicillium oxalicum. Biotechnol Biofuels 9:216PubMedGoogle Scholar
  60. Xue JL, Zhao S, Liang RM, Yin X, Jiang SX, Su LH, Yang Q, Duan CJ, Liu JL, Feng JX (2016) A biotechnological process efficiently co-produces two high value-added products, glucose and xylooligosaccharides, from sugarcane bagasse. Bioresour Technol 204:130–138PubMedGoogle Scholar
  61. Yan YS, Zhao S, Liao LS, He QP, Xiong YR, Wang L, Li CX, Feng JX (2017) Transcriptomic profiling and genetic analyses reveal novel key regulators of cellulase and xylanase gene expression in Penicillium oxalicum. Biotechnol Biofuels 10:279PubMedPubMedCentralGoogle Scholar
  62. Yang Q, Gao Y, Huang YP, Xu QS, Luo XM, Liu JL, Feng JX (2015) Identification of three important amino acid residues of xylanase AfxynA from Aspergillus fumigatus for enzyme activity and formation of xylobiose as the major product. Process Biochem 50:571–581Google Scholar
  63. Zhang Z, Liu JL, Lan JY, Duan CJ, Ma QS, Feng JX (2014) Predominance of Trichoderma and Penicillium in cellulolytic aerobic filamentous fungi from subtropical and tropical forests in China, and their use in finding highly efficient beta-glucosidase. Biotechnol Biofuels 7:107Google Scholar

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources, Guangxi Research Center for Microbial and Enzyme Engineering Technology, College of Life Science and TechnologyGuangxi UniversityNanningPeople’s Republic of China

Personalised recommendations