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Applied and Environmental Microbiology, July 2000, p. 3016-3023, Vol. 66, No. 7
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Studies on the Production of Fungal Peroxidases in Aspergillus niger

Ana Conesa, Cees A. M. J. J. van den Hondel, and Peter J. Punt*

Department of Molecular Genetics and Gene Technology, TNO Nutrition and Food Research Institute, 3704 HE Zeist, The Netherlands

Received 31 August 1999/Accepted 17 April 2000

To get insight into the limiting factors existing for the efficient production of fungal peroxidase in filamentous fungi, the expression of the Phanerochaete chrysosporium lignin peroxidase H8 (lipA) and manganese peroxidase (MnP) H4 (mnp1) genes in Aspergillus niger has been studied. For this purpose, a protease-deficient A. niger strain and different expression cassettes have been used. Northern blotting experiments indicated high steady-state mRNA levels for the recombinant genes. Manganese peroxidase was secreted into the culture medium as an active protein. The recombinant protein showed specific activity and a spectrum profile similar to those of the native enzyme, was correctly processed at its N terminus, and had a slightly lower mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Recombinant MnP production could be increased up to 100 mg/liter upon hemoglobin supplementation of the culture medium. Lignin peroxidase was also secreted into the extracellular medium, although the protein was not active, presumably due to incorrect processing of the secreted enzyme. Expression of the lipA and mnp1 genes fused to the A. niger glucoamylase gene did not result in improved production yields.

* Corresponding author. Mailing address: Department of Molecular Genetics and Gene Technology, TNO Nutrition and Food Research Institute, P.O. Box 360, 3700 AJ Zeist, The Netherlands. Phone: 31 30 6944463. Fax: 31 30 6944466. E-mail: P.Punt{at}voeding.tno.nl.

Applied and Environmental Microbiology, July 2000, p. 3016-3023, Vol. 66, No. 7
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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