Advertisement

Cellulose

, Volume 25, Issue 3, pp 1865–1881 | Cite as

Cellulose fiber size defines efficiency of enzymatic hydrolysis and impacts degree of synergy between endo- and exoglucanases

  • Vanessa O. A. Pellegrini
  • Amanda Bernardes
  • Camila A. Rezende
  • Igor Polikarpov
Original Paper
  • 201 Downloads

Abstract

An interplay between cellulases is fundamental in biomass saccharification. Here, the synergistic action of Trichoderma harzianum Cel7A and Cel7B on two cellulosic substrates: bacterial cellulose (BC) and a much more heterogeneous filter paper (FP) was investigated by determining their saccharification yields and by analyzing both the released soluble products and the insoluble reducing ends formed in the process. Furthermore, morphological changes of the substrates were evaluated using scanning electron microscopy. Glycoside hydrolase family 7 (GH7) enzymes introduce uniform changes in BC, whereas in FP they preferentially consume thin microfibrils rather than thicker paper fibers. Thus, the size effect, which leads to a smaller surface area per unit of substrate mass for thicker fibers, seems to play a crucial role in higher enzymatic hydrolysis efficiency of BC as compared to FP. These results demonstrate that the morphology-dependent effects could be essential for the industrial breakdown of cellulose-rich plant biomass.

Keywords

Trichoderma harzianum Cel7A Cel7B Synergism Scanning electron microscopy 

Notes

Acknowledgments

We would like to acknowledge support of the Brazilian funding agencies FAPESP via grants 2010/18773-8, 2012/22802-9, 2015/13684-0 and 2016/13602-7 and CNPq via grants 472523/2013-9 and 405191/2015-4. The electron microscopy work has been performed using the Quanta 650 microscope at LME/LNNano/CNPEM, Campinas.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.

Supplementary material

10570_2018_1700_MOESM1_ESM.docx (428 kb)
Supplementary material 1 (DOCX 427 kb)

References

  1. Arantes V, Saddler J (2010) Access to cellulose limits the efficiency of enzymatic hydrolysis: the role of amorphogenesis. Biotechnol Biofuels 3:4.  https://doi.org/10.1186/1754-6834-3-4 CrossRefGoogle Scholar
  2. Badino SF, Christensen SJ, Kari J, Windahl MS, Hvidt S, Borch K, Westh P (2017) Exo–exo synergy between Cel6A and Cel7A from Hypocrea jecorina: role of carbohydrate binding module and the endo-lytic character of the enzymes. Biotechnol Bioeng 114:1639–1647.  https://doi.org/10.1002/bit.26276 CrossRefGoogle Scholar
  3. Bernardinelli OD, Lima MA, Rezende CA, Polikarpov I, deAzevedo ER (2015) Quantitative 13C MultiCP solid-state NMR as a tool for evaluation of cellulose crystallinity index measured directly inside sugarcane biomass. Biotechnol Biofuels 8:110.  https://doi.org/10.1186/s13068-015-0292-1 CrossRefGoogle Scholar
  4. Boisset C, Fraschini C, Schülein M, Henrissat B, Chanzy H (2000) Imaging the enzymatic digestion of bacterial cellulose ribbons reveals the endo character of the cellobiohydrolase Cel6A from Humicola insolens and its mode of synergy with cellobiohydrolase Cel7A. Appl Environ Microbiol 66:1444–1452.  https://doi.org/10.1128/aem.66.4.1444-1452.2000 CrossRefGoogle Scholar
  5. Boisset C, Pétrequin C, Chanzy H, Henrissat B, Schülein M (2001) Optimized mixtures of recombinant Humicola insolens cellulases for the biodegradation of crystalline cellulose. Biotechnol Bioeng 72:339–345.  https://doi.org/10.1002/1097-0290(20010205)72:3<339::AID-BIT11>3.0.CO;2-%23
  6. Chawla PR, Bajaj IB, Survase SA, Singhal RS (2009) Microbial cellulose: fermentative production and applications. Food Technol Biotechnol 47:107–124Google Scholar
  7. Colussi F, Serpa V, Delabona PS, Manzine LR, Voltatodio ML, Alves R, Mello BL, Pereira N Jr, Farinas CS, Golubev AM, Santos MA, Polikarpov I (2011) Purification, and biochemical and biophysical characterization of cellobiohydrolase I from Trichoderma harzianum IOC 3844. J Microbiol Biotechnol 21:808–817.  https://doi.org/10.4014/jmb.1010.10037 CrossRefGoogle Scholar
  8. Cruys-Bagger N, Tatsumi H, Ren GR, Borch K, Westh P (2013) Transient kinetics and rate-limiting steps for the processive cellobiohydrolase Cel7A: effects of substrate structure and carbohydrate binding domain. Biochemistry 52:8938–8948.  https://doi.org/10.1021/bi401210n CrossRefGoogle Scholar
  9. Ganner T, Bubner P, Eibinger M, Mayrhofer C, Plank H, Nidetzky B (2012) Dissecting and reconstructing synergism: in situ visualization of cooperativity among cellulases. J Biol Chem 287:43215–43222.  https://doi.org/10.1074/jbc.M112.419952 CrossRefGoogle Scholar
  10. Goodwin JW (2004) Colloids and Interfaces with surfactants and polymers—an introduction. Wiley, London.  https://doi.org/10.1002/0470093919 CrossRefGoogle Scholar
  11. Harris P, Welner D, McFarland K, Re E, Navarro Poulsen J, Brown K, Salbo R, Ding H, Vlasenko E, Merino S (2010) Stimulation of lignocellulosic biomass hydrolysis by proteins of glycoside hydrolase family 61: structure and function of a large, enigmatic family. Biochemistry 49:3305–3316.  https://doi.org/10.1021/bi100009p CrossRefGoogle Scholar
  12. Hemsworth GR, Johnston EM, Davies GJ, Walton PH (2015) Lytic polysaccharide monooxygenases in biomass conversion. Trends Biotechnol 33:747–761.  https://doi.org/10.1016/j.tibtech.2015.09.006 CrossRefGoogle Scholar
  13. Henrissat B, Driguez H, Viet C, Schulein M (1985) Synergism of cellulases from Trichoderma reesei in the degradation of cellulose. Nat Biotechnol 3:722–726.  https://doi.org/10.1038/nbt0885-722 CrossRefGoogle Scholar
  14. Horn SJ, Vaaje-Kolstad G, Westereng B, Eijsink VGH (2012) Novel enzymes for the degradation of cellulose. Biotechnol Biofuels 5:1–13.  https://doi.org/10.1186/1754-6834-5-45 CrossRefGoogle Scholar
  15. Hunter RJ (2001) Foundations of colloid science, 2nd edn. Oxford University Press, OxfordGoogle Scholar
  16. Igarashi K, Uchihashi T, Koivula A, Wada M, Kimura S, Okamoto T, Penttilä M, Ando T, Samejima M (2011) Traffic jams reduce hydrolytic efficiency of cellulase on cellulose surface. Science 333:1279–1282.  https://doi.org/10.1126/science.l208386 CrossRefGoogle Scholar
  17. Jalak J, Kurašin M, Teugjas H, Väljamäe P (2012) Endo-exo synergism in cellulose hydrolysis revisited. J Biol Chem 287:28802–28815.  https://doi.org/10.1074/jbc.M112.381624 CrossRefGoogle Scholar
  18. Jeoh T, Wilson DB, Walker LP (2006) Effect of cellulase mole fraction and cellulose recalcitrance on synergism in cellulose hydrolysis and binding. Biotechnol Prog 22:270–277.  https://doi.org/10.1021/bp050266f CrossRefGoogle Scholar
  19. Kari J, Olsen J, Borch K, Cruys-Bagger N, Jensen K, Westh P (2014) Kinetics of cellobiohydrolase (Cel7A) variants with lowered substrate affinity. J Biol Chem 47:32459–32468.  https://doi.org/10.1074/jbc.M114.604264 CrossRefGoogle Scholar
  20. Katzen F (2007) Gateway® recombinational cloning: a biological operating system. Exp Opin Drug Discov 2:571–589.  https://doi.org/10.1517/17460441.2.4.571 CrossRefGoogle Scholar
  21. Kostylev M, Wilson DB (2012) Synergistic interactions in cellulose hydrolysis. Biofuels 3:61–70.  https://doi.org/10.4155/bfs.11.150 CrossRefGoogle Scholar
  22. Kostylev M, Wilson D (2014) A distinct model of synergism between a processive endocellulase (TfCel9A) and an exocellulase (TfCel48A) from Thermobifida fusca. Appl Environ Microbiol 80:339–344.  https://doi.org/10.1128/AEM.02706-13 CrossRefGoogle Scholar
  23. Kurasin M, Väljamäe P (2011) Processivity of cellobiohydrolases is limited by the substrate. J Biol Chem 286:169–177.  https://doi.org/10.1074/jbc.M110.161059 CrossRefGoogle Scholar
  24. Kuusk S, Sørlie M, Väljamäe P (2015) The predominant molecular state of bound enzyme determines the strength and type of product inhibition in the hydrolysis of recalcitrant polysaccharides by processive enzymes. J Biol Chem 290:11678–11691.  https://doi.org/10.1074/jbc.M114.635631 CrossRefGoogle Scholar
  25. Li Y, Irwin DC, Wilson DB (2007) Processivity, substrate binding, and mechanism of cellulose hydrolysis by Thermobifida fusca Cel9A. Appl Environ Microbiol 73:3165–3172.  https://doi.org/10.1128/AEM.02960-06 CrossRefGoogle Scholar
  26. Luterbacher JS, Moran-Mirabal JM, Burkholder EW, Walker LP (2015) Modeling enzymatic hydrolysis of lignocellulosic substrates using confocal fluorescence microscopy I: filter paper cellulose. Biotechnol Bioeng 112:21–31.  https://doi.org/10.1002/bit.25329 CrossRefGoogle Scholar
  27. Miller G (1959) Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal Chem 31:426–428.  https://doi.org/10.1021/ac60147a030 CrossRefGoogle Scholar
  28. Nidetzky B, Steiner W, Hayn M, Claeyssens M (1994) Cellulose hydrolysis by the cellulases from Trichoderma reesei: a new model for synergistic interaction. Biochem J 298:705–710.  https://doi.org/10.1042/bj29880705 CrossRefGoogle Scholar
  29. Park S, Baker JO, Himmel ME, Parilla PA, Johnson DK (2010) Cellulose crystallinity index: measurement techniques and their impact on interpreting cellulase performance. Biotechnol Biofuels 3:10.  https://doi.org/10.1186/1754-6834-3-10 CrossRefGoogle Scholar
  30. Payne CM, Knott BC, Mayes HB, Hansson H, Himmel ME, Sandgren M, Ståhlberg J, Beckham GT (2015) Fungal cellulases. ‎Chem Rev 115:1308–1448.  https://doi.org/10.1021/cr500351c CrossRefGoogle Scholar
  31. Pellegrini VO, Serpa VI, Godoy AS, Camilo CM, Bernardes A, Rezende CA, Junior NP, Franco Cairo JP, Squina FM, Polikarpov I (2015) Recombinant Trichoderma harzianum endoglucanase I (Cel7B) is a highly acidic and promiscuous carbohydrate-active enzyme. Appl Microbiol Biotechnol 99:9591–9604.  https://doi.org/10.1007/s00253-015-6772-1 CrossRefGoogle Scholar
  32. Reese ET, Siu RGH, Levinson HS (1950) The biological degradation of soluble cellulose derivatives and its relationship to the mechanism of cellulose hydrolysis. J Bacteriol 59:485–497Google Scholar
  33. Rezende CA, Lima MA, Maziero P, de Azevedo ER, Garcia W, Polikarpov I (2011) Chemical and morphological characterization of sugarcane bagasse submitted to a delignification process for enhanced enzymatic digestibility. Biotechnol Biofuels 4:54.  https://doi.org/10.1186/1754-6834-4-54 CrossRefGoogle Scholar
  34. Schneider CA, Rasband WS, Eliceiri KW (2012) NIH Image to ImageJ: 25 years of image analysis. Nat Methods 9:671–675.  https://doi.org/10.1038/nmeth.2089 CrossRefGoogle Scholar
  35. Schramm M, Hestrin S (1954) Synthesis of cellulose by Acetobacter xylinum. 1. Micromethod for the determination of celluloses. Biochem J 56:163–166.  https://doi.org/10.1042/bj0560163 CrossRefGoogle Scholar
  36. Serpa VI, Polikarpov I (2011) Enzymes in bioenergy. In: Goldman MSBH (ed) Routes to cellulosic ethanol, 1st edn. Springer, New York, pp 97–113.  https://doi.org/10.1007/978-0-387-92740-4_7 CrossRefGoogle Scholar
  37. Silveira MH, Aguiar RS, Siika-aho M, Ramos LP (2014) Assessment of the enzymatic hydrolysis profile of cellulosic substrates based on reducing sugar release. Bioresour Technol 151:392–396.  https://doi.org/10.1016/j.biortech.2013.09.135 CrossRefGoogle Scholar
  38. Ståhlberg J, Johansson G, Pettersson G (1993) Trichoderma reesei has no true exo-cellulase: all intact and truncated cellulases produce new reducing end groups on cellulose. Biochim Biophys Acta 1157:107–113.  https://doi.org/10.1016/0304-4165(93)90085-M CrossRefGoogle Scholar
  39. Storms R, Zheng Y, Li H, Sillaots S, Martinez-Perez A, Tsang A (2005) Plasmid vectors for protein production, gene expression and molecular manipulations in Aspergillus niger. Plasmid 53:191–204.  https://doi.org/10.1016/j.plasmid.2004.10.001 CrossRefGoogle Scholar
  40. Textor LC, Colussi F, Silveira RL, Serpa V, de Mello BL, Muniz JRC, Squina FM, Pereira N, Skaf MS, Polikarpov I (2013) Joint X-ray crystallographic and molecular dynamics study of cellobiohydrolase I from Trichoderma harzianum: deciphering the structural features of cellobiohydrolase catalytic activity. FEBS J 280:56–69.  https://doi.org/10.1111/febs.12049 CrossRefGoogle Scholar
  41. Tomazini A, Dolce LG, de Oliveira Neto M, Polikarpov I (2015) Xanthomonas campestris expansin-like X domain is a structurally disordered beta-sheet macromolecule capable of synergistically enhancing enzymatic efficiency of cellulose hydrolysis. Biotechnol Lett 37:2419–2426.  https://doi.org/10.1007/s10529-015-1927-9 CrossRefGoogle Scholar
  42. Vaaje-Kolstad G, Westereng B, Horn SJ, Liu ZL, Zhai H, Sørlie M, Eijsink VGH (2010) An oxidative enzyme boosting the enzymatic conversion of recalcitrant polysaccharides. Science 330:219–222.  https://doi.org/10.1126/science.1192231 CrossRefGoogle Scholar
  43. Väljamäe P, Sild V, Nutt A, Pettersson G, Johansson G (1999) Acid hydrolysis of bacterial cellulose reveals different modes of synergistic action between cellobiohydrolase I and endoglucanase I. Eur J Biochem 266:327–334.  https://doi.org/10.1046/j.1432-1327.1999.00853.x CrossRefGoogle Scholar
  44. Ververis C, Georghiou K, Christodoulakis N, Santas P, Santas R (2004) Fiber dimensions, lignin and cellulose content of various plant materials and their suitability for paper production. Ind Crop Prod 19:245–254.  https://doi.org/10.1016/j.indcrop.2003.10.006 CrossRefGoogle Scholar
  45. Wang T, Park YB, Caporini MA, Rosay M, Zhong L, Cosgrove DJ, Hong M (2013) Sensitivity-enhanced solid-state NMR detection of expansin’s target in plant cell walls. PNAS 110:16444–16449.  https://doi.org/10.1073/pnas.1316290110 CrossRefGoogle Scholar
  46. Watson DL, Wilson DB, Walker LP (2002) Synergism in binary mixtures of Thermobifida fusca cellulases Cel6B, Cel9A, and Cel5A on BMCC and Avicel. Appl Biochem Biotechnol 101:97–111.  https://doi.org/10.1385/ABAB:101:2:097 CrossRefGoogle Scholar
  47. Wilson DB, Kostylev M (2012) Cellulase processivity. Methods in molecular biology. Methods Mol Biol 908:93–99.  https://doi.org/10.1007/978-1-61779-956-3_9 Google Scholar
  48. Woodward J, Hayes MK, Lee NE (1988) Hydrolysis of cellulose by saturating and non-saturating concentrations of cellulase: implications for synergism. Nat Biotechnol 6:301–304.  https://doi.org/10.1038/nbt0388-301 CrossRefGoogle Scholar
  49. Zhang YHP, Lynd LR (2004) Toward an aggregated understanding of enzymatic hydrolysis of cellulose: noncomplexed cellulase systems. Biotechnol Bioeng 88:797–824.  https://doi.org/10.1002/bit.20282 CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V., part of Springer Nature 2018

Authors and Affiliations

  1. 1.Department of Physics and Interdisciplinary Science, São Carlos Institute of PhysicsUniversity of São PauloSão CarlosBrazil
  2. 2.Institute of ChemistryUniversity of CampinasCampinasBrazil

Personalised recommendations