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Production of Cellobiose Dehydrogenase from a Newly Isolated White Rot Fungus Termitomyces sp. OE147

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Abstract

Class I cellobiose dehydrogenases (CDHs) are extracellular hemoflavo enzymes produced at low levels by the Basidiomycetes (white rot fungi). In presence of suitable electron acceptors, e.g., cytochrome c, 2,6-dichlorophenol-indophenol, or metal ions, it oxidizes cellobiose to cellobionolactone. A stringent requirement for disaccharides makes CDH also useful for conversion of lactose to lactobionic acid, an important ingredient in pharma and detergent industry. In this work, class I CDH was produced using a newly identified white rot fungus Termitomyces sp. OE147. Four media were evaluated for CDH production, and maximum enzyme activity of 0.92 international unit (IU)/ml was obtained on Ludwig medium under submerged conditions. Statistical optimization of N source, which had significant effect on CDH production, using Box–Behnken design followed by optimization of inoculum size and age resulted in an increase in activity to 2.9 IU/ml and a productivity of ~25 IU/l/h. The nearly purified CDH exhibited high activity of 26.4 IU/mg protein on lactose indicating this enzyme to be useful for lactobionic acid synthesis. Some of the internal peptide sequences bore 100 % homology to the CDH produced in Myceliophthora thermophila. The fungal isolate was amenable to scale up, and an overall productivity of ~18 IU/l/h was obtained at 14-l level.

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References

  1. Cameron, M. D., & Aust, S. D. (2001). Cellobiose dehydrogenase—an extracellular fungal flavocytochrome. Enzyme and Microbial Technology, 28, 129–138.

    Article  CAS  Google Scholar 

  2. Zamocky, M., Ludwig, R., Peterbauer, C., Hallberg, B., Divne, C., & Nicholls, P. (2006). Cellobiose dehydrogenase—a flavocytochrome from wood—degrading, phytopathogenic and saprotropic fungi. Current Protein and Peptide Science, 7, 255–280.

    Article  CAS  Google Scholar 

  3. Bao, W., Usha, S. N., & Renganathan, V. (1993). Purification and characterization of cellobiose dehydrogenase, a novel extracellular hemoflavoenzyme from the white-rot fungus Phanerochaete chrysosporium. Archives of Biochemistry and Biophysics, 300, 705–713.

    Article  CAS  Google Scholar 

  4. Xu, F., Golightly, E. J., Duke, K. R., Lassen, S. F., Knusen, B., Christensen, S., et al. (2001). Humicola insolens cellobiose dehydrogenase: cloning, redox chemistry, and “logic gate”-like dual functionality. Enzyme and Microbial Technology, 28, 744–753.

    Article  CAS  Google Scholar 

  5. Baminger, U., Subramaniam, S. S., Renganathan, V., & Haltrich, D. (2001). Purification and characterization of cellobiose dehydrogenase from the plant pathogen Sclerotium (Athelia) rolfsii. Applied and Environmental Microbiology, 67, 1766–1774.

    Article  CAS  Google Scholar 

  6. Henriksson, G., Johansson, G., & Petterson, G. (2000). A critical review of cellobiose dehydrogenase. Journal of Biotechnology, 78, 93–113.

    Article  CAS  Google Scholar 

  7. Henriksson, G., Zhang, L. J., Ljungquist, P., Reitberger, T., Petterson, G., & Johansson, G. (2000). Is cellobiose dehydrogenase from Phanerochaete chrysosporium a lignin degrading enzyme? Biochimica et Biophysica Acta, 1480, 83–91.

    Article  CAS  Google Scholar 

  8. Dumonceaux, T., Bartholomew, K., Laleanu, L., Charles, T., & Archibald, F. (2001). Cellobiose dehydrogenase is essential for wood invasion and non-essential for Kraft pulp delignification by Trametes versicolor. Enzyme and Microbial Technology, 29, 478–489.

    Article  CAS  Google Scholar 

  9. Dhariwal, A., Mavrov, V., & Scroeder, I. (2006). Production of lactobionic acid with process integrated electrochemical enzyme regeneration and optimisation of process variables using response surface methods (RSM). Journal of Molecular Catalysis B: Enzymatic, 42, 64–69.

    Article  CAS  Google Scholar 

  10. Barbara, A., Green, R. P., Richard, H., Wildnauer, P. D., & Brenda, L. E. Lactobionic acid- a novel polyhydroxy bionic acid for skincare. Princeton: Neostrata Company, Inc.

  11. Fang, J., Huang, F., & Gao, P. (1999). Optimization of cellobiose dehydrogenase production by Schizophyllum commune and effect of the enzyme on kraft pulp bleaching by ligninases. Process Biochemistry, 34, 957–961.

    Article  CAS  Google Scholar 

  12. Harreither, W., Sygmund, C., Dunhofen, E., Vicuna, R., Haltrich, D., & Ludwig, R. (2009). Celloboise dehydrogenase from the ligninolytic basidiomycetes Ceriporiopsis subvermispora. Applied and Environmental Microbiology, 75, 2750–2757.

    Article  CAS  Google Scholar 

  13. Li, B., Rotsaert, A. J. F., Gold, M. H., & Renganathan, V. (2000). Homologous expression of recombinant cellobiose dehydrogenase in Phanerochaete chrysosporium. Biochemical and Biophysical Research Communications, 270, 141–146.

    Article  CAS  Google Scholar 

  14. Yoshida, M., Ohira, T., Igarashi, K., Nagasawa, H., Aida, K., Hallberg, B. M., et al. (2001). Production and characterization of recombinant Phanerochaete chrysosporium cellobiose dehydrogenase in the methylotrophic yeast Pichia pastoris. Bioscience, Biotechnology, and Biochemistry, 65, 2050–2057.

    Article  CAS  Google Scholar 

  15. Stapleton, P. C., O’Brien, M. M., O’Callaghan, J., & Dobson, A. D. W. (2004). Molecular cloning of the cellobiose dehydrogenase from Trametes versicolor and expression in Pichia pastoris. Enzyme and Microbial Technology, 34, 55–63.

    Article  CAS  Google Scholar 

  16. Zamocky, M., Schümann, C., Sygmund, C., Dobson, A. D., Ludwig, R., Haltrich, D., et al. (2008). Cloning, sequence analysis and heterologous expression in Pichia pastoris of a gene encoding a thermostable cellobiose dehydrogenase from Myriococcum thermophilum. Protein Expression and Purification, 59, 258–265.

    Article  CAS  Google Scholar 

  17. Zhang, R., Fan, Z., & Kasuga, T. (2011). Expression of cellobiose dehydrogenase from Neurospora crassa in Pichia pastoris and its purification and characterization. Protein Expression and Purification, 75, 63–69.

    Article  CAS  Google Scholar 

  18. Ludwig, R., Salamon, A., Varga, J., Zamocky, M., Peterbauer, C. K., & Kulbe, K. D. (2004). Characterization of cellobiose dehydrogenases from the white-rot fungi Trametes pubescens and Trametes villosa. Applied Microbiology and Biotechnology, 64, 213–222.

    Article  CAS  Google Scholar 

  19. Connick, D., Bouquelet, J., & Dumoitiers, D. (2000). Industrial media and fermentation processes for improved growth and protease production by Tetrahymena thermophila. Journal of Industrial Microbiology and Biotechnology, 24, 285–290.

    Article  Google Scholar 

  20. Saha, T., Ghosh, D., Mukherjee, S., Bose, S., & Mukherjee, M. (2008). Cellobiose dehydrogenase production by the mycelial culture of the mushroom Termitomyces clypeatus. Process Biochemistry, 48, 634–641.

    Article  Google Scholar 

  21. Nystrom, J. M., & DiLuca, P. H. Enhanced production of Trichoderma cellulase on high levels of cellulose in submerged culture. IIT Delhi Symposium 1977.

  22. Biswas, R., Sahai, V., Mishra, S., & Bisaria, V. S. (2010). Bioprocess strategies for enhanced production of xylanase by Melanocarpus albomyces IITD3A on agro—residual extract. Journal of Bioscience and Bioengineering, 110, 702–708.

    Article  CAS  Google Scholar 

  23. Box, G. E. P., & Behnken, D. W. (1960). Some new three level design for the study of quantitative variables. Technometrics, 2, 455–475.

    Article  Google Scholar 

  24. Baminger, U., Nidetzky, B., Kulbe, K. D., & Haltrich, D. (1999). A simple assay for measuring cellobiose dehydrogenase activity in the presence of laccase. Journal of Microbiological Methods, 35, 253–259.

    Article  CAS  Google Scholar 

  25. Canevascini, G., Borer, P., & Dreyer, J. L. (1991). Cellobiose dehydrogenase of Sporotricum (Chrysosporium) thermophile. European Journal of Biochemistry, 198, 43–52.

    Article  CAS  Google Scholar 

  26. Ghosh, T. K. (1987). Measurement of cellulase activities. Pure and Applied Chemistry, 59, 257–268.

    Google Scholar 

  27. Rogalski, J., Szczodrak, J., & Janusz, G. (2006). Manganese peroxidase production in submerged cultures by free and immobilized mycelia of Nematoloma frowardii. Bioresource Technology, 97, 469–476.

    Article  CAS  Google Scholar 

  28. Arora, D. S., & Gill, P. K. (2001). Comparison of two assay procedures for lignin peroxidase. Enzyme and Microbial Technology, 28, 602–605.

    Article  CAS  Google Scholar 

  29. Temp, U., & Eggert, C. (1999). Novel interaction between laccase and cellobiose dehydrogenase during pigment synthesis in the white rot fungus Pycnoporus cinnabarinus. Applied and Environmental Microbiology, 65, 389–395.

    CAS  Google Scholar 

  30. Meyer, S. P., & Ahearn, D. G. (1977). Extracellular proteolysis by Candida lipolytica. Mycologia, 69, 646–651.

    Article  Google Scholar 

  31. Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein–dye binding. Analytical Biochemistry, 72, 248–254.

    Article  CAS  Google Scholar 

  32. Song, A. R., Tian, X. M., Feng, G. F., & Zhang, Y. Q. (2001). A study of the utilization of Pleurotus eryngii with different carbon and nitrogen sources. Acta Edulis Fungi, 8, 10–14.

    Google Scholar 

  33. Qinnghe, C., Xiaoyu, Y., Tiangui, N., Cheng, J., & Qiugang, M. (2004). The screening of culture conditions and properties of xylanase by white-rot fungus Pleurotus ostreatus. Process Biochemistry, 39, 1561–1566.

    Article  Google Scholar 

  34. Gruno, M., Valjamae, P., Pettersson, G., & Johansson, G. (2004). Inhibition of the Trichoderma ressei cellulases by cellobiose is strongly dependent on the nature of the substrate. Biotechnology and Bioengineering, 86, 503–511.

    Article  CAS  Google Scholar 

  35. Subramaniam, S. S., Nagalla, S. R., & Reganathan, V. (1999). Cloning and characterization of a thermostable cellobiose dehydrogenase from Sporotrichum thermophile. Archives of Biochemistry and Biophysics, 365, 223–230.

    Article  CAS  Google Scholar 

  36. Yoshida, M., Ohira, T., Igarashi, K., Nagasawa, H., & Samejima, M. (2002). Molecular cloning and characterization of a cDNA encoding cellobiose dehydrogenase from the wood-rotting fungus Grifola frondosa. FEMS Microbiology Letters, 217, 225–230.

    Article  CAS  Google Scholar 

  37. Bailey, J. E., & Ollis, D. F. (1986) Biochemical engineering fundamentals, 2nd Ed. New York: McGraw Hill, Inc.

  38. Sikyta, B. (1983). Development of microbial processes. In B. Sikyta & M. Prause (Eds.), Methods in industrial microbiology (pp. 250–274). Chichester: Ellis Horwood Ltd.

    Google Scholar 

  39. Archibald, F. S., Bourbonnais, R., Jurasek, L., Paice, M. G., & Reid, I. D. (1997). Kraft pulp bleaching and delignification by Trametes versicolor. Journal of Biotechnology, 53, 215–236.

    Article  CAS  Google Scholar 

  40. Kudriavtseva, O. A., Dunaevskii, L. E., Kamzolkina, O. V., & Belozerskii, M. A. (2008). Proteolytic enzymes of the fungi: extracellular proteases of xylotrophic basidiomycetes. Mikrobiologiia, 77, 725–737.

    CAS  Google Scholar 

  41. Sabotic, J., Trcek, T., Popovic, T., & Brzin, J. (2007). Basidiomycets harbor a hidden treasure of proteolytic activity. Journal of Biotechnology, 128, 297–307.

    Article  CAS  Google Scholar 

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Acknowledgments

Authors gratefully acknowledge the financial assistance received from the Department of Biotechnology (Government of India) for carrying out this study. Further, we are thankful to Dr. R. C. Upadhyay from the National Research Centre for Mushroom (NRCM) Solan, India, for providing the cultures. The authors would also like to acknowledge the help of Mr. M.V.R.K. Sarma for the analysis of the statistical data.

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Authors and Affiliations

  1. Department of Biochemical Engineering and Biotechnology, Indian Institute of Technology Delhi, New Delhi, 110016, India

    Gupteshwar Gupta, Rishabh Gangwar, Ashwani Gautam, Lalit Kumar, Vikram Sahai & Saroj Mishra

  2. JAIV Technologies Pvt., Ltd., Sector 10 Dwarka, New Delhi, 110075, India

    Anuj Dhariwal

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  1. Gupteshwar Gupta
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  2. Rishabh Gangwar
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Correspondence to Saroj Mishra.

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Gupta, G., Gangwar, R., Gautam, A. et al. Production of Cellobiose Dehydrogenase from a Newly Isolated White Rot Fungus Termitomyces sp. OE147. Appl Biochem Biotechnol 173, 2099–2115 (2014). https://doi.org/10.1007/s12010-014-1010-3

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