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Applied Microbiology and Biotechnology

, Volume 100, Issue 6, pp 2907–2915 | Cite as

Clostridium thermocellum releases coumaric acid during degradation of untreated grasses by the action of an unknown enzyme

  • Christopher D. Herring
  • Philip G. Thorne
  • Lee R. Lynd
Bioenergy and biofuels

Abstract

Clostridium thermocellum is an anaerobic thermophile with the ability to digest lignocellulosic biomass that has not been pretreated with high temperatures. Thermophilic anaerobes have previously been shown to more readily degrade grasses than wood. Part of the explanation for this may be the presence of relatively large amounts of coumaric acid in grasses, with linkages to both hemicellulose and lignin. We found that C. thermocellum and cell-free cellulase preparations both release coumaric acid from bagasse and switchgrass. Cellulase preparations from a mutant strain lacking the scaffoldin cipA still showed activity, though diminished. Deletion of all three proteins in C. thermocellum with ferulic acid esterase domains, either singly or in combination, did not eliminate the activity. Further work will be needed to identify the novel enzyme(s) responsible for the release of coumaric acid from grasses and to determine whether these enzymes are important factors of microbial biomass degradation.

Keywords

Coumaric acid esterase Cellulosic ethanol Cellulolytic microorganisms Analytical chemistry 

Notes

Acknowledgments

The authors wish to thank Julie Paye for helpful discussion, William Kenealy, and Abigail Foster for providing C. thermocellum cellulase and Dan Olson for providing strain M1668.

Compliance with ethical standards

Funding

This research was funded by Mascoma Corporation and the BioEnergy Science Center, a US Department of Energy Bioenergy Research Center supported by the Office of Biological and Environmental Research in the DOE Office of Science. This manuscript has been authored by a contractor of the US Government under contract DE-AC05-00OR22725.

Conflict of interest

The authors of this study have been employed by Mascoma Corporation and/or Enchi Corporation, which have held a financial interest in technology related to C. thermocellum and T. saccharolyticum.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

References

  1. Akin DE (2007) Grass lignocellulose: strategies to overcome recalcitrance. Appl Biochem Biotech 137–140:3–15Google Scholar
  2. Argyros DA, Tripathi SA, Barrett TF, Rogers SR, Feinberg LF, Olson DG, Foden JM, Miller BB, Lynd LR, Hogsett DA, Caiazza NC (2011) High ethanol titers from cellulose by using metabolically engineered thermophilic, anaerobic microbes. Appl Environ Microbiol 77:8288–8294. doi: 10.1128/AEM.00646-11 PubMedCentralCrossRefPubMedGoogle Scholar
  3. Barthelmebs L, Divies C, Cavin JF (2000) Knockout of the p-coumarate decarboxylase gene from Lactobacillus plantarum reveals the existence of two other inducible enzymatic activities involved in phenolic acid metabolism. Appl Environ Microbiol 66:3368–3375. doi: 10.1128/AEM.66.8.3368-3375.2000 PubMedCentralCrossRefPubMedGoogle Scholar
  4. Basen M, Rhaesa AM, Kataeva I, Prybol CJ, Scott IM, Poole FL, Adams MWW (2014) Degradation of high loads of crystalline cellulose and of unpretreated plant biomass by the thermophilic bacterium Caldicellulosiruptor bescii. Bioresour Technol 152:384–392. doi: 10.1016/j.biortech.2013.11.024 CrossRefPubMedGoogle Scholar
  5. Baskaran S, Ahn HJ, Lynd LR (1995) Investigation of the ethanol tolerance of Clostridium thermosaccharolyticum in continuous culture. Biotechnol Prog 11:276–281. doi: 10.1021/bp00033a006 CrossRefGoogle Scholar
  6. Benoit I, Danchin EGJ, Bleichrodt RJ, de Vries RP (2008) Biotechnological applications and potential of fungal feruloyl esterases based on prevalence, classification and biochemical diversity. Biotechnol Lett 30:387–396CrossRefPubMedGoogle Scholar
  7. Blum DL, Kataeva IA, Li XL, Ljungdahl LG (2000) Feruloyl esterase activity of the Clostridium thermocellum cellulosome can be attributed to previously unknown domains of XynY and XynZ. J Bacteriol 182:1346–1351. doi: 10.1128/JB.182.5.1346-1351.2000 PubMedCentralCrossRefPubMedGoogle Scholar
  8. Bugg T, Ahmad M, Hardiman EM, Rahmanpour R (2011) Pathways for degradation of lignin in bacteria and fungi. Nat Prod Rep 28:1883. doi: 10.1039/c1np00042j CrossRefPubMedGoogle Scholar
  9. Chung D, Cha M, Guss AM, Westpheling J (2014) Direct conversion of plant biomass to ethanol by engineered Caldicellulosiruptor bescii. Proc Natl Acad Sci U S A 111:8931–6. doi: 10.1073/pnas.1402210111 PubMedCentralCrossRefPubMedGoogle Scholar
  10. Crepin VF, Faulds CB, Connerton IF (2004) Functional classification of the microbial feruloyl esterases. Appl Microbiol Biotechnol 63:647–652CrossRefPubMedGoogle Scholar
  11. Gaillard BD, Richards GN (1975) Presence of soluble lignin-carbohydrate complexes in the bovine rumen. Carbohydr Res 42:135–145. doi: 10.1016/S0008-6215(00)84106-3 CrossRefPubMedGoogle Scholar
  12. Gibson DG (2011) Enzymatic assembly of overlapping DNA fragments. Methods Enzymol 498:349–361. doi: 10.1016/B978-0-12-385120-8.00015-2 CrossRefPubMedGoogle Scholar
  13. Gnansounou E, Dauriat A (2010) Techno-economic analysis of lignocellulosic ethanol: a review. Bioresour Technol 101:4980–4991. doi: 10.1016/j.biortech.2010.02.009 CrossRefPubMedGoogle Scholar
  14. Grabber JH, Hatfield RD, Ralph J (1998) Diferulate cross-links impede the enzymatic degradation of non-lignified maize walls. J Sci Food Agric 77:193–200. doi: 10.1002/(SICI)1097-0010(199806)77:2<193::AID-JSFA25>3.0.CO;2-A CrossRefGoogle Scholar
  15. Hartley RD, Harris PJ (1981) Phenolic constituents of the cell walls of dicotyledons. Biochem Syst Ecol 9:189–203CrossRefGoogle Scholar
  16. Hogsett DA (1995) Cellulose hydrolysis and fermentation by Clostridium thermocellum for the production of ethanol. PhD Thesis, Dartmouth College, Hanover, NH, USAGoogle Scholar
  17. Izquierdo JA, Pattathil S, Guseva A, Hahn MG, Lynd LR (2014) Comparative analysis of the ability of Clostridium clariflavum strains and Clostridium thermocellum to utilize hemicellulose and unpretreated plant material. Biotechnol Biofuels 7:1–8. doi: 10.1186/s13068-014-0136-4 CrossRefGoogle Scholar
  18. Kaneko T, Thi TH, Shi DJ, Akashi M (2006) Environmentally degradable, high-performance thermoplastics from phenolic phytomonomers. Nat Mater 5:966–970. doi: 10.1038/nmat1778 CrossRefPubMedGoogle Scholar
  19. Kataeva I, Foston MB, Yang S-J, Pattathil S, Biswal AK, Ii FLP, Basen M, Rhaesa AM, Thomas TP, Azadi P, Olman V, Saffold TD, Mohler KE, Lewis DL, Doeppke C, Zeng Y, Tschaplinski TJ, York WS, Davis M, Mohnen D, Xu Y, Ragauskas AJ, Ding S-Y, Kelly RM, Hahn MG, Adams MWW (2013) Carbohydrate and lignin are simultaneously solubilized from unpretreated switchgrass by microbial action at high temperature. Energy Environ Sci 6:2186–2195. doi: 10.1039/c3ee40932e CrossRefGoogle Scholar
  20. Koseki T, Fushinobu S, Ardiansyah, Shirakawa H, Komai M (2009) Occurrence, properties, and applications of feruloyl esterases. Appl Microbiol Biotechnol 84:803–810CrossRefPubMedGoogle Scholar
  21. Kumar D, Murthy GS (2011) Impact of pretreatment and downstream processing technologies on economics and energy in cellulosic ethanol production. Biotechnol Biofuels 4:27. doi: 10.1186/1754-6834-4-27 PubMedCentralCrossRefPubMedGoogle Scholar
  22. Lam TBT, Kadoya K, Iiyama K (2001) Bonding of hydroxycinnamic acids to lignin: ferulic and p-coumaric acids are predominantly linked at the benzyl position of lignin, not the β-position, in grass cell walls. Phytochemistry 57:987–992. doi: 10.1016/S0031-9422(01)00052-8
  23. Lam TBT, Iiyama K, Stone BA (2003) Hot alkali-labile linkages in the walls of the forage grass Phalaris aquatica and Lolium perenne and their relation to in vitro wall digestibility. Phytochemistry 64:603–607. doi: 10.1016/S0031-9422(03)00301-7 CrossRefPubMedGoogle Scholar
  24. Lee JM, Venditti RA, Jameel H, Kenealy WR (2011) Detoxification of woody hydrolyzates with activated carbon for bioconversion to ethanol by the thermophilic anaerobic bacterium Thermoanaerobacterium saccharolyticum. Biomass Bioenergy 35:626–636. doi: 10.1016/j.biombioe.2010.10.021 CrossRefGoogle Scholar
  25. Lynd LR, Elamder RT, Wyman CE (1996) Likely features and costs of mature biomass ethanol technology. Appl Biochem Biotechnol 57–58:741–761CrossRefGoogle Scholar
  26. Lynd LR, Weimer PJ, van Zyl WH, Pretorius IS (2002) Microbial cellulose utilization: fundamentals and biotechnology. Microbiol Mol Biol Rev 66:506–577. doi: 10.1128/MMBR.66.3.506-577.2002 PubMedCentralCrossRefPubMedGoogle Scholar
  27. Masarin F, Gurpilhares DB, Baffa DC, Barbosa MH, Carvalho W, Ferraz A, Milagres AM (2011) Chemical composition and enzymatic digestibility of sugarcane clones selected for varied lignin content. Biotechnol Biofuels 4:55. doi: 10.1186/1754-6834-4-55 PubMedCentralCrossRefPubMedGoogle Scholar
  28. Mathew S, Abraham TE (2004) Ferulic acid: an antioxidant found naturally in plant cell walls and feruloyl esterases involved in its release and their applications. Crit Rev Biotechnol 24:59–83. doi: 10.1080/07388550490491467 CrossRefPubMedGoogle Scholar
  29. Morag E, Yaron S, Lamed R, Kenig R, Shoham Y, Bayer EA (1996) Dissociation of the cellulosome of Clostridium thermocellum under nondenaturing conditions. J Biotechnol 51:235–242. doi: 10.1016/S0168-1656(96)01601-X CrossRefGoogle Scholar
  30. Neilson MJ, Richards GN (1978) The fate of the soluble lignin-carbohydrate complex produced in the bovine rumen. J Sci Food Agric 29:513–519CrossRefPubMedGoogle Scholar
  31. Olson DG, Lynd LR (2012) Transformation of Clostridium thermocellum by electroporation. Methods Enzymol 510:317–330. doi: 10.1016/B978-0-12-415931-0.00017-3 CrossRefPubMedGoogle Scholar
  32. Olson DG, Giannone RJ, Hettich RL, Lynd LR (2013) Role of the CipA scaffoldin protein in cellulose solubilization, as determined by targeted gene deletion and complementation in Clostridium thermocellum. J Bacteriol 195:733–739. doi: 10.1128/JB.02014-12 PubMedCentralCrossRefPubMedGoogle Scholar
  33. Paliwal R, Rawat AP, Rawat M, Rai JPN (2012) Bioligninolysis: recent updates for biotechnological solution. Appl Biochem Biotechnol 167:1865–1889. doi: 10.1007/s12010-012-9735-3 CrossRefPubMedGoogle Scholar
  34. Paye J, Guseva A, Hammer S, Gjersing E, Davis M, Davison B, Olstad J, Donohoe B, Nguyen T, Wyman C, Pattathil S, Hahn M, Lynd L (2015) Biological lignocellulose solubilization: comparative evaluation of biocatalysts and enhancement via cotreatment. Biotechnol Biofuels (in press)Google Scholar
  35. Pu Y, Hu F, Huang F, Davison BH, Ragauskas AJ (2013) Assessing the molecular structure basis for biomass recalcitrance during dilute acid and hydrothermal pretreatments. Biotechnol Biofuels 6:15. doi: 10.1186/1754-6834-6-15 PubMedCentralCrossRefPubMedGoogle Scholar
  36. Rakotoarivonina H, Hermant B, Chabbert B, Touzel JP, Remond C (2011) A thermostable feruloyl-esterase from the hemicellulolytic bacterium Thermobacillus xylanilyticus releases phenolic acids from non-pretreated plant cell walls. Appl Microbiol Biotechnol 90:541–552. doi: 10.1007/s00253-011-3103-z CrossRefPubMedGoogle Scholar
  37. Saulnier L, Vigouroux J, Thibault JF (1995) Isolation and partial characterization of feruloylated oligosaccharides from maize bran. Carbohydr Res 272:241–253. doi: 10.1016/0008-6215(95)00053-V CrossRefPubMedGoogle Scholar
  38. Scalbert A (1985) Ether linkage between phenolic acids and lignin fractions from wheat straw. Phytochemistry 24:1359–1362CrossRefGoogle Scholar
  39. Shin SY, Han NS, Park YC, Kim MD, Seo JH (2011) Production of resveratrol from p-coumaric acid in recombinant Saccharomyces cerevisiae expressing 4-coumarate:coenzyme A ligase and stilbene synthase genes. Enzyme Microb Technol 48:48–53. doi: 10.1016/j.enzmictec.2010.09.004 CrossRefPubMedGoogle Scholar
  40. Siqueira G, Milagres AM, Carvalho W, Koch G, Ferraz A (2011) Topochemical distribution of lignin and hydroxycinnamic acids in sugar-cane cell walls and its correlation with the enzymatic hydrolysis of polysaccharides. Biotechnol Biofuels 4:7. doi: 10.1186/1754-6834-4-7 PubMedCentralCrossRefPubMedGoogle Scholar
  41. Sun RC, Sun XF, Zhang SH (2001) Quantitative determination of hydroxycinnamic acids in wheat, rice, rye, and barley straws, maize stems, oil palm frond fiber, and fast-growing poplar wood. J Agric Food Chem 49:5122–5129. doi: 10.1021/jf010500r CrossRefPubMedGoogle Scholar
  42. Talluri S, Raj SM, Christopher LP (2013) Consolidated bioprocessing of untreated switchgrass to hydrogen by the extreme thermophile Caldicellulosiruptor saccharolyticus DSM 8903. Bioresour Technol 139:272–279. doi: 10.1016/j.biortech.2013.04.005 CrossRefPubMedGoogle Scholar
  43. Tan K, Chang C, Cuff M, Osipiuk J, Landorf E, Mack JC, Zerbs S, Joachimiak A, Collart FR (2013) Structural and functional characterization of solute binding proteins for aromatic compounds derived from lignin: p-coumaric acid and related aromatic acids. Proteins Struct Funct Bioinform 81:1709–1726. doi: 10.1002/prot.24305 CrossRefGoogle Scholar
  44. Wong DWS (2006) Feruloyl esterase: a key enzyme in biomass degradation. Appl Biochem Biotechnol 133:87–112. doi: 10.1385/ABAB:133:2:87 CrossRefPubMedGoogle Scholar
  45. Yang B, Wyman CE (2008) Pretreatment: the key to unlocking low-cost cellulosic ethanol. Biofuels Bioprod Biorefining 2:26–40CrossRefGoogle Scholar
  46. Yang SJ, Kataeva I, Hamilton-Brehm SD, Engle NL, Tschaplinski TJ, Doeppke C, Davis M, Westpheling J, Adams MWW (2009) Efficient degradation of lignocellulosic plant biomass, without pretreatment, by the thermophilic anaerobe “Anaerocellum thermophilum” DSM 6725. Appl Environ Microbiol 75:4762–4769. doi: 10.1128/AEM.00236-09 PubMedCentralCrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Christopher D. Herring
    • 1
    • 2
    • 3
  • Philip G. Thorne
    • 2
  • Lee R. Lynd
    • 1
    • 2
    • 3
  1. 1.Thayer School of Engineering, Dartmouth CollegeHanoverUSA
  2. 2.Mascoma CorporationLebanonUSA
  3. 3.Enchi CorporationWalthamUSA

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