Title
Natural and recombinant fungal laccases for paper pulp bleaching
Author
Sigoillot, C.
Record, E.
Belle, V.
Robert, J.L.
Levasseur, A.
Punt, P.J.
van den Hondel, C.A.M.J.J.
Fournel, A.
Sigoillot, J.C.
Asther, M.
TNO Voeding
Publication year
2004
Abstract
Three laccases, a natural form and two recombinant forms obtained from two different expression hosts, were characterized and compared for paper pulp bleaching. Laccase from Pycnoporus cinnabarinus, a well known lignolytic fungus, was selected as a reference for this study. The corresponding recombinant laccases were produced in Aspergillus oryzae and A. niger hosts using the lacI gene from P. cinnabarinus to develop a production process without using the expensive laccase inducers required by the native source. In flasks, production of recombinant enzymes by Aspergilli strains gave yields close to 80 mg l -1. Each protein was purified to homogeneity and characterized, demonstrating that the three hosts produced proteins with similar physico-chemical properties, including electron paramagnetic resonance spectra and N-terminal sequences. However, the recombinant laccases have higher Michaelian (Km) constants, suggesting a decrease in substrate/enzyme affinity in comparison with the natural enzyme. Moreover, the natural laccase exhibited a higher redox potential (around 810 mV), compared with A. niger (760 mV) and A. oryzae (735 mV). Treatment of wheat straw Kraft pulp using laccases expressed in P. cinnabarinus or A. niger with 1-hydroxybenzotriazole as redox mediator achieved a delignification close to 75%, whereas the recombinant laccase from A. oryzae was not able to delignify pulp. These results were confirmed by thioacidolysis. Kinetic and redox potential data and pulp bleaching results were consistent, suggesting that the three enzymes are different and each fungal strain introduces differences during protein processing (folding and/or glycosylation).
Subject
Biology
Biotechnology
fungal enzyme
laccase
recombinant enzyme
fungus
amino terminal sequence
article
Aspergillus niger
Aspergillus oryzae
biotechnology
bleaching
controlled study
delignification
enzyme analysis
enzyme induction
enzyme substrate
Michaelis constant
nonhuman
nucleotide sequence
oxidation reduction reaction
physical chemistry
protein synthesis
pulp processing
Aspergillus niger
Aspergillus oryzae
Biotechnology
Cloning, Molecular
Genes, Fungal
Industrial Microbiology
Laccase
Lignin
Oxidation-Reduction
Paper
Polyporaceae
Recombinant Proteins
Substrate Specificity
Aspergillus
Aspergillus niger
Aspergillus oryzae
Fungi
Pycnoporus cinnabarinus
Triticum aestivum
To reference this document use:
http://resolver.tudelft.nl/uuid:ebfed3f6-2800-40fc-8d4d-7e30c6e3887a
DOI
https://doi.org/10.1007/s00253-003-1468-3
TNO identifier
237697
ISSN
0175-7598
Source
Applied Microbiology and Biotechnology, 64 (3), 346-352
Document type
article