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Heterologous Production of a Ligninolytic Enzyme: Expression of the Phlebia Radiata Laccase Gene in Trichoderma Reesei

Bio/Technology volume 9, pages 987–990 (1991)Cite this article

Abstract

We have expressed a gene encoding the ligninolytic laccase enzyme of the white–rot fungus Phlebia radiata in the soft rot fungus Trichoderma reesei under the promoter of the major cellulase gene (cbh1). Constructions from the laccase cDNA or the chromosomal gene were equivalent in expression, yielding 20 mg/l of secreted active laccase in small–scale fermentations. The recombinant enzyme has a similar molecular weight, antigenic properties and specific activity as the Phlebia laccase and is analogous to the Phlebia enzyme in removing monomeric lignin–related compounds from residual lignin of the pulping process, thus demonstrating its potential for industrial applications.

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References

  1. Kirk, T.K. and Farrell, R.L. 1987. Enzymatic “combustion”: The microbial degradation of lignin. Ann. Rev. Microbiol. 41: 465–505.

    Article  CAS  Google Scholar 

  2. Niku-Paavola, M-L., Karhunen, E., Salola, P. and Raunio, V. 1988. Ligninolytic enzymes of the white-rot fungus Phlebia radiata. Biochem. J. 254: 877–884.

    Article  CAS  Google Scholar 

  3. Karhunen, E., Kantelinen, A. and Niku-Paavola, M.-L. 1990. Mn-dependent peroxidase from the lignin-degrading white-rot fungus Phlebia radiata. Arch. Biochem. Biophys. 279: 25–31.

    Article  CAS  Google Scholar 

  4. Karhunen, E., Niku-Paavola, M.-L., Viikari, L., Haltia, T., van de Meer, R.A. and Duine, J.A. 1990. A novel combination of prosthetic groups in a fungal laccase: PQQ and two copper atoms. FEBS Letts. 267: 6–8.

    Article  CAS  Google Scholar 

  5. Saloheimo, M., Barajas, V., Niku-Paavola, M.-L. and Knowles, J.K.C. 1989. A lignin peroxidase-encoding cDNA from the white-rot fungus Phlebia radiata: characterization and expression in Trichoderma reesei. Gene 85: 343–351.

    Article  CAS  Google Scholar 

  6. Saloheimo, M., Niku-Paavola, M.-L. and Knowles, J.K.C. 1991. Isolation and structural analysis of the laccase gene from the lignin-degrading fungus Phlebia radiata. J. Gen. Microbiol. In press.

    Google Scholar 

  7. Saunders, G., Picknett, T.M., Tuite, M.F. and Ward, M. 1989. Heterologous gene expression in filamentous fungi. Trends in Biotechnol. 7: 283–287.

    Article  CAS  Google Scholar 

  8. Harkki, A., Uusitalo, J., Bailey, M., Penttilä, M. and Knowles, J.K.C. 1989. A novel fungal expression system: secretion of active calf chymosin from the filamentous fungus Trichoderma reesei. Bio/Technology 7: 596–603.

    CAS  Google Scholar 

  9. Uusitalo, J.M., Nevalainen, K.M.H., Harkki, A.M., Knowles, J.K.C. and Penttilä, M.E. 1991. Enzyme production by recombinant Trichoderma reesei strains. J. Biotechnol. 17: 35–50.

    Article  CAS  Google Scholar 

  10. Mattern, I.E., Punt, P.J., Unkles, S., Pouvels, P.H. and van den Hondel, C.A.M.J.J. 1987. Transfomation of Aspergillus oryzae. In: Abstracts of the 19th Lunteren Lectures on Molecular Genetics of Yeasts and Filamentous Fungi and its Impact on Biotechnology, p. 34. Lunteren, The Netherlands.

    Google Scholar 

  11. Hynes, M.J., Corrick, C.M. and King, J.A. 1983. Isolation of genomic clones containing the amdS gene of Aspergillus nidulans and their use in the analysis of structural and regulatory mutations. Mol. Cell. Biol. 3: 1430–1439.

    Article  CAS  Google Scholar 

  12. Ballance, D.J. 1986. Sequences important for gene expression in filamentous fungi. Yeast 2: 229–236.

    Article  CAS  Google Scholar 

  13. Penttilä, M., Lehtovaara, P., Nevalainen, H., Bhikhabhai, R. and Knowles, J.K.C. 1986. Homology between cellulase genes of Trichoderma reesei: complete nucleotide sequence of the endoglucanase I gene. Gene 45: 253–263.

    Article  Google Scholar 

  14. Kuan, I. and Tien, M. 1989. Phosphorylation of lignin peroxidases from Phanerochaete chrysosporium. J. Biol. Chem. 264: 20350–20355.

    CAS  PubMed  Google Scholar 

  15. Davis, S. and Burns, R.G. 1990. Decolorization of phenolic effluents by soluble and immobilized phenol oxidases. Appl. Microbiol. Biotechnol. 32: 721–726.

    Article  CAS  Google Scholar 

  16. Farrell, R., Gelep, P., Anilionis, A., Javaherian, K., Maione, T.E., Rushe, J.R., Sadownick, B.A. and Jackson, J.A. 1987. Recombinant ligninase. European Patent Application No. 87810516.2.

    Google Scholar 

  17. Kojima, Y., Tsukuda, Y., Kawai, Y., Tsukamoto, A., Sugiura, J. and Sakaino, M., 1990. Cloning, sequence analysis, and expression of ligninolytic phenoloxidase genes of the white-rot basidiomycete Coriolus hirsutus. J. Biol. Chem. 265: 15224–15230.

    CAS  PubMed  Google Scholar 

  18. Niku-Paavola, M.-L., Karhunen, E., Kantelinen, A., Viikari, L., Lundell, T. and Hatakka, A. 1990. The effect of culture conditions on the production of lignin modifying enzymes by the white-rot fungus Phlebia radiata. J. Biotechnol. 13: 211–221.

    Article  CAS  Google Scholar 

  19. Ward, M., Wilson, L.J., Kodama, K.H., Rey, M.W. and Berka, R.M. 1990. Improved production of chymosin in Aspergillus by expression as a glucoamylase-chymosin fusion. Bio/Technology 8: 435–440.

    CAS  PubMed  Google Scholar 

  20. Lamsa, M. and Bloebaum, P. 1990. Mutation and screening to increase chymosin yield in a genetically-engineered strain of Aspergillus awamori. J. Industr. Microbiol. 5: 229–238.

    Article  CAS  Google Scholar 

  21. Sheir-Neiss, G. and Montenecourt, B.S. 1984. Characterization of the secreted cellulases of Trichoderma reesei wild type and mutants during controlled fermentations. Appl. Microbiol. Biotechnol. 20: 46–53.

    Article  CAS  Google Scholar 

  22. Yannish-Perron, C., Vieira, J. and Messing, J. 1985. Improved M13 phage cloning vectors and host strains: nucleotide sequence of the M13mpl8 and pUC19 vectors. Gene 33: 103–119.

    Article  Google Scholar 

  23. Penttilä, M., Nevalainen, H., Rättö, M., Salminen, E. and Knowles, J. 1987. A versatile transformation system for the cellulolytic filamentous fungus Trichoderma reesei.Gene 61: 155–164.

    Article  Google Scholar 

  24. Raeder, U. and Broda, P. 1985. Rapid preparation of DNA from filamentous fungi. Lett. Appl. Microbiol. 1: 17–20.

    Article  CAS  Google Scholar 

  25. Chirgwin, J.M., Przybyla, A.E., MacDonald, R.J. and Rutter, W.J. 1979. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochem. J. 18: 5294–5299.

    Article  CAS  Google Scholar 

  26. Bailey, M.J. and Ojamo, H. 1990. Selective concentration of polygalacturonase and beta-D-glucosidase of Aspergillus niger culture filtrate using mineral absorbents. Bioseparation 1: 133–139.

    CAS  Google Scholar 

  27. Bradford, M.M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding.Anal. Biochem. 72: 248–254.

    Article  CAS  Google Scholar 

  28. Niku-Paavola, M.-L., Raaska, L. and Itävaara, M. 1990. Detection of white-rot fungi by a nontoxic stain. Mycol. Res. 94: 27–31.

    Article  Google Scholar 

  29. Alen, R. and Vikkula, A. 1989. Formation of lignin monomers during alkaline delignification of softwood. Holzforschung 43: 397–400.

    Article  CAS  Google Scholar 

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  1. Markku Saloheimo: Corresponding author.

Authors and Affiliations

  1. VTT, Biotechnical Laboratory, P.O. Box 202, SF-02151, Espoo, Finland

    Markku Saloheimo & Marja-Leena Niku-Paavola

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  1. Markku Saloheimo
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  2. Marja-Leena Niku-Paavola
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Saloheimo, M., Niku-Paavola, ML. Heterologous Production of a Ligninolytic Enzyme: Expression of the Phlebia Radiata Laccase Gene in Trichoderma Reesei. Nat Biotechnol 9, 987–990 (1991). https://doi.org/10.1038/nbt1091-987

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