Applied Microbiology and Biotechnology

, Volume 32, Issue 4, pp 436–442 | Cite as

Screening of basidiomycetes for lignin peroxidase genes using a DNA probe

  • Yoshio Kimura
  • Yasuhiko Asada
  • Masaaki Kuwahara


Basidiomycetes were screened for lignin peroxidase (LPO) genes using a DNA probe prepared from the LPO restriction fragment ofPhanerochaete chrysosporium. Southern blot analysis showed restriction fragments of chromosomal DNA ofBjerkandera adusta andCoriolus consors hybridized with the probe.Bjerkandera adusta produced LPO in a glucose-peptone medium. Ion-exchange chromatography showed that this fungus produced multiple molecular forms of LPO. One of the enzymes, LPO-2, was purified and characterized. The molecular weight of LPO-2 was 41000 with a pI of 4.2. Spectral analysis demonstrated that LPO-2 is a haem protein. The enzyme cleaved lignin model dimers mainly at the Cα-Cβ position of the side chain. The LPO-2 exhibited close similarity to LPOs ofP. chrysosporium with respect to their basic properties.


Lignin Southern Blot Restriction Fragment Southern Blot Analysis Molecular Form 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Asada Y, Kimura Y, Kuwahara M, Tsukamoto A, Koide K, Oka A, Takanami M (1988) Cloning and sequencing of a ligninase gene from a lignin-degrading basidiomycete,Phanerochaete chrysosporium. Appl Microbiol Biotechnol 29:469–473Google Scholar
  2. Boer HA de, Zhang YZ, Collins C, Reddy CA (1987) Analysis of nucleotide sequences of two ligninase cDNAs from a white-rot filamentous fungus,Phanerochaete chrysosporium. Gene 60:93–102Google Scholar
  3. Bradford MM (1976) A rapid and sensitive method for the quantification of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254Google Scholar
  4. Buswell JA, Mollet B, Odier E (1984) Ligninolytic enzyme production byPhanerochaete chrysosporium under conditions of nitrogen sufficiency. FEMS Microbiol Lett 25:295–299Google Scholar
  5. Dodson PJ, Evans CS, Harvey PJ, Palmer JM (1987) Production and properties of an extracellular peroxidase from Coriolus versicolor which catalyses Cα-Cβ cleavage in a lignin model compound. FEMS Microbiol Lett 42:17–22Google Scholar
  6. Faison BD, Kirk TK (1985) Factors involved in the regulation of a ligninase activity inPhanerochaete chrysosporium. Appl Environ Microbiol 49:299–304Google Scholar
  7. Gold MH, Kuwahara M, Chin AA, Glenn JK (1984) Purification and characterization of an extracellular H2O2-requir-ing diarylpropane oxygenase from the white rot basidiomycete,Phanerochaete chrysosporium. Arch Biochem Biophys 234:353–362Google Scholar
  8. Gold MH, Wariishi H, Valli K. (1989) Extracellular peroxidases involved in lignin degradation by the white rot basidiomycetePhanerochaete chrysosporium. In: Whitaker JR, Sonnet PE (eds) Biocatalysis in agricultural biotechnology, ACS Symposium Series no. 389. American Chemical Society, Washington DC, USA, pp 127–140Google Scholar
  9. Jeffries TW, Choi S, Kirk TK (1981) Nutritional regulation of lignin. degradation byPhanerochaete chrysosporium. Appl Environ Microbiol 42:290–296Google Scholar
  10. Kirk TK, Schultz E, Connors WJ, Lorenz LF, Zeikus JG (1978) Influence of culture parameters on lignin metabolism byPhanerochaete chrysosporium. Arch Microbiol 117:277–285Google Scholar
  11. Kirk K, Croan S, Tien M, Murtagh KE, Farrell RL (1986) Production of multiple ligninases byPhanerochaete chrysosporium: effect of selected growth conditions and use of a mutant strain. Enzyme Microb Technol 8:27–32Google Scholar
  12. Kirk TK, Kersten PJ, Mozuch MD, Kalyanaraman B (1986) Ligninase ofPhanerochaete chrysosporium. Mechanism of its degradation of the non-phenolic arylglycerol β-aryl ether substructure of lignin. Biochem J 236:279–287Google Scholar
  13. Kirk TK, Farrell RL (1987) Enzymatic “combustion”: the microbial degradation of lignin. Ann Rev Microbiol 41:465–505Google Scholar
  14. Kos A, Kuijvenhoven J, Wernars K, Bos CJ, Broek HWJ van den, Pouwels PH, Hondel CAMJJ van den (1985) Isolation and characterization of theAspergillus niger trpC gene. Gene 39:231–238Google Scholar
  15. Kuwahara M, Glenn JK, Morgan MA, Gold MH (1984) Separation and characterization of two extracellular H2O2-de-pendent oxidases from ligninolytic cultures ofPhanerochaete chrysosporium. FEBS Lett 169:247–250Google Scholar
  16. Kuwahara M, Asada Y, Kimura Y, Aokage M (1987) Isolation of mutants of a lignin-degrading basidiomycete,Phanerochaete chrysosporium, that produce lignin peroxidase in high nitrogen cultures. Mokuzai Gakkaishi 33:821–823Google Scholar
  17. Leisola MSA, Kozulic B, Meussdoerffer F, Fiechter A (1987) Homology among multiple extracellular peroxidases fromPhanerochaete chrysosporium. J Biol Chem 262:419–424Google Scholar
  18. Niku-Paavola M-L, Karhunen E, Salola P, Raunio V (1988) Ligninolytic enzymes of the white-rot fungusPhlebia radiata. Biochem J 254:877–884Google Scholar
  19. Paszczynski A, Huynh V-B, Crawford R (1986) Comparison of ligninase-I and peroxidase-M2 from the white-rot fungusPhanerochaete chrysosporium. Arch Biochem Biophys 244:750–765Google Scholar
  20. Smith TL, Schalch H, Gaskell J, Covert S, Cullen D (1988) Nucleotide sequence of a ligninase gene fromPhanerochaete chrysosporium. Nucleic Acids Res 16:1219Google Scholar
  21. Tien M, Kirk TK (1984) Lignin-degrading enzyme fromPhanerochaete chrysosporium: purification, characterization, and catalytic properties of a unique H2O2-requiring oxygenase. Proc Natl Acad Sci USA 81:2280–2284Google Scholar
  22. Tien M, Tu C-PD (1987) Cloning and sequencing of a cDNA for a ligninase fromPhanerochaete chrysosporium. Nature 326:520–523Google Scholar
  23. Upshall A, Gilbert T, Saari G, O'Hara PJ, Weglenski P, Berse B, Miller K, Timberlake WE (1986) Molecular analysis of theargB gene ofAspergillus nidulans. Mol Gen Genet 204:349–354Google Scholar
  24. Waldner R, Leisola MSA, Fiechter A (1988) Comparison of ligninolytic activities of selected white-rot fungi. Appl Microbiol Biotechnol 29:400–407Google Scholar
  25. Walther I, Kalin M, Reiser J, Suter F, Fritsche B, Saloheimo M, Leisola M, Teeri T, Knowles JKC, Fiechter A (1988) Molecular analysis of a Phanerochaete chrysosporium lignin peroxidase gene. Gene 70:127–137Google Scholar
  26. Ward M, Wilson LJ, Carmona CL, Turner G (1988) The oliC3 gene ofAspergillus niger: isolation, sequence and use as a selectable marker for transformation. Curr Genet 14:37–42Google Scholar

Copyright information

© Springer-Verlag 1990

Authors and Affiliations

  • Yoshio Kimura
    • 1
  • Yasuhiko Asada
    • 1
  • Masaaki Kuwahara
    • 1
  1. 1.Department of Bioresource ScienceKagawa UniversityKagawaJapan

Personalised recommendations