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Biofuels pp 213-231 | Cite as

Cellulase Assays

Protocol
First Online:
Part of the Methods in Molecular Biology book series (MIMB, volume 581)

Summary

Cellulose is a heterogeneous polysaccharide, and its enzymatic hydrolysis requires endoglucanase, exoglucanase (cellobiohydrolase), and β-glucosidase to work together. We summarize the most commonly used assays for individual enzymes and cellulase mixture.

Key words

β-Glucosidase Cellobiase Cellobiohydrolase Cellulose Cellulase assay Endoglucanase Sugar assay 
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Notes

Acknowledgments

This work was made possible in part by support from the Biological Systems Engineering Department of Virginia Polytechnic Institute and State University and USDA-CSREES (2006–38909–03484) and DOE BioEnergy Science Center (BESC).

References

  1. 1.
    Lynd LR, Weimer PJ, van Zyl WH, and Pretorius IS (2002) Microbial cellulose utilization: fundamentals and biotechnology. Microbiol. Mol. Biol. Rev. 66:506–577.CrossRefGoogle Scholar
  2. 2.
    Zhang Y-HP, Himmel M, and Mielenz JR (2006) Outlook for cellulase improvement: screening and selection strategies. Biotechnol. Adv. 24(5):452–481.CrossRefGoogle Scholar
  3. 3.
    Zhang Y-HP and Lynd LR (2004) Toward an aggregated understanding of enzymatic hydrolysis of cellulose: noncomplexed cellulase systems. Biotechnol. Bioeng. 88:797–824.CrossRefGoogle Scholar
  4. 4.
    Zhang Y-HP (2008) Reviving the carbohydrate economy via multi-product biorefineries. J. Ind. Microbiol. Biotechnol. 35(5):367–375.CrossRefGoogle Scholar
  5. 5.
    Wood TM and Bhat KM (1988) Methods for measuring cellulase activities. Methods Enzymol. 160:87–117.CrossRefGoogle Scholar
  6. 6.
    Ghose TK (1987) Measurement of cellulase activities. Pure Appl. Chem. 59:257–268.CrossRefGoogle Scholar
  7. 7.
    Zhang Y-HP and Lynd LR (2006) A functionally-based model for hydrolysis of cellulose by fungal cellulase. Biotechnol. Bioeng. 94:888–898.CrossRefGoogle Scholar
  8. 8.
    Hong J, Ye X, and Zhang Y-HP (2007) Quantitative determination of cellulose accessibility to cellulase based on adsorption of a nonhydrolytic fusion protein containing CBM and GFP with its applications. Langmuir 23(25):12535–12540.CrossRefGoogle Scholar
  9. 9.
    Morag E, Bayer EA, and Lamed RL (1992) Affinity digestion for the near-total recovery of purified cellulosome from Clostridium thermocellum. Enzyme Microb. Technol. 14:289–292.CrossRefGoogle Scholar
  10. 10.
    Coward-Kelly G, Aiello-Mazzari C, Kim S, Granda C, and Holtzapple M (2003) Suggested improvements to the standard filter paper assay used to measure cellulase activity. Biotechnol. Bioeng. 82:745–749.CrossRefGoogle Scholar
  11. 11.
    Johnson EA, Sakajoh M, Halliwell G, Madia A, and Demain AL (1982) Saccharification of complex cellulosic substrates by the cellulase system from Clostridium thermocellum. Appl. Environ. Microbiol. 43:1125–1132.Google Scholar
  12. 12.
    Zhang Y-HP and Lynd LR (2003) Quantification of cell and cellulase mass concentrations during anaerobic cellulose fermentation: development of an ELISA-based method with application to Clostridium thermocellum batch cultures. Anal. Chem. 75:219–227.CrossRefGoogle Scholar
  13. 13.
    Zhang Y-HP and Lynd LR (2005) Regulation of cellulase synthesis in batch and continuous cultures of Clostridium thermocellum. J. Bacteriol. 187:99–106.CrossRefGoogle Scholar
  14. 14.
    Zhang Y-HP and Lynd LR (2004) Kinetics and relative importance of phosphorolytic and hydrolytic cleavage of cellodextrins and cellobiose in cell extracts of Clostridium thermocellum. Appl. Environ. Microbiol. 70:1563–1569.CrossRefGoogle Scholar
  15. 15.
    Hong J, Ye X, Wang Y, and Zhang Y-HP (2008) Bioseparation of recombinant cellulose binding module-protein by affinity adsorption on an ultra-high-capacity cellulosic adsorbent. Anal. Chem. Acta 621:193–199.CrossRefGoogle Scholar
  16. 16.
    Zhang Y-HP and Lynd LR (2005) Determination of the number-average degree of polymerization of cellodextrins and cellulose with application to enzymatic hydrolysis. Biomacromolecules 6:1510–1515.CrossRefGoogle Scholar
  17. 17.
    Zhang Y-HP, Cui J-B, Lynd LR, and Kuang LR (2006) A transition from cellulose swelling to cellulose dissolution by o-phosphoric acid: evidences from enzymatic hydrolysis and supramolecular structure. Biomacromolecules 7(2):644–648.CrossRefGoogle Scholar
  18. 18.
    Miller GL (1959) Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal. Chem. 31:426–428.CrossRefGoogle Scholar
  19. 19.
    Zhang Y-HP, Schell DJ, and McMillan JD (2007) Methodological analysis for determination of enzymatic digestibility of cellulosic materials. Biotechnol. Bioeng. 96(1):188–194.CrossRefGoogle Scholar
  20. 20.
    Kongruang S, Han MJ, Breton CIG, and Penner MH (2004) Quantitative analysis of cellulose-reducing ends. Appl. Biochem. Biotechnol. 113–116:213–231.CrossRefGoogle Scholar

Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  1. 1.Biological Systems Engineering DepartmentVirginia Polytechnic Institute and State UniversityBlacksburgUSA

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