1. Natural and recombinant fungal laccases for paper pulp bleaching
- Author
-
André Fournel, Peter J. Punt, Anthony Levasseur, Michèle Asther, Eric Record, C. Sigoillot, J. L. Robert, Valérie Belle, C.A.M.J.J. van den Hondel, Jean-Claude Sigoillot, Unité mixte de recherche de biotechnologie des champignons filamenteux, Université de Provence - Aix-Marseille 1-Institut National de la Recherche Agronomique (INRA)-Université de la Méditerranée - Aix-Marseille 2, Centre National de la Recherche Scientifique (CNRS), Netherlands Organisation for Applied Scientific Research, TNO Voeding, and Université de la Méditerranée - Aix-Marseille 2-Institut National de la Recherche Agronomique (INRA)-Université de Provence - Aix-Marseille 1
- Subjects
enzyme induction ,protein synthesis ,Aspergillus oryzae ,Applied Microbiology and Biotechnology ,Lignin ,law.invention ,laccase ,Substrate Specificity ,Polyporaceae ,chemistry.chemical_compound ,law ,Cloning, Molecular ,recombinant enzyme ,0303 health sciences ,delignification ,biology ,Pulp (paper) ,fungus ,Fungal genetics ,article ,General Medicine ,pulp processing ,fungal enzyme ,Recombinant Proteins ,Aspergillus ,[SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology ,Biochemistry ,Recombinant DNA ,Aspergillus niger ,Michaelis constant ,Oxidation-Reduction ,biotechnology ,Biotechnology ,Paper ,Genes, Fungal ,Triticum aestivum ,engineering.material ,03 medical and health sciences ,Industrial Microbiology ,physical chemistry ,controlled study ,Biology ,enzyme analysis ,030304 developmental biology ,enzyme substrate ,Laccase ,nonhuman ,030306 microbiology ,Fungi ,nucleotide sequence ,Pycnoporus cinnabarinus ,bleaching ,biology.organism_classification ,PULPE DE PAPIER ,chemistry ,engineering ,amino terminal sequence ,RECOMBINANT PROTEIN ,oxidation reduction reaction - Abstract
International audience; 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 (K m) 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).
- Published
- 2003