Highly Efficient Production of Laccase by the Basidiomycete Pycnoporus cinnabarinus |
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Authors: | Alexandra M C R Alves Eric Record Anne Lomascolo Karin Scholtmeijer Marcel Asther Joseph G H Wessels and Han A B Wsten |
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Institution: | Alexandra M. C. R. Alves, Eric Record, Anne Lomascolo, Karin Scholtmeijer, Marcel Asther, Joseph G. H. Wessels, and Han A. B. Wösten |
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Abstract: | An efficient transformation and expression system was developed for the industrially relevant basidiomycete Pycnoporus cinnabarinus. This was used to transform a laccase-deficient monokaryotic strain with the homologous lac1 laccase gene placed under the regulation of its own promoter or that of the SC3 hydrophobin gene or the glyceraldehyde-3-phosphate dehydrogenase (GPD) gene of Schizophyllum commune. SC3-driven expression resulted in a maximal laccase activity of 107 nkat ml−1 in liquid shaken cultures. This value was about 1.4 and 1.6 times higher in the cases of the GPD and lac1 promoters, respectively. lac1-driven expression strongly increased when 25 g of ethanol liter−1 was added to the medium. Accordingly, laccase activity increased to 1,223 nkat ml−1. These findings agree with the fact that ethanol induces laccase gene expression in some fungi. Remarkably, lac1 mRNA accumulation and laccase activity also strongly increased in the presence of 25 g of ethanol liter−1 when lac1 was expressed behind the SC3 or GPD promoter. In the latter case, a maximal laccase activity of 1,393 nkat ml−1 (i.e., 360 mg liter−1) was obtained. Laccase production was further increased in transformants expressing lac1 behind its own promoter or that of GPD by growth in the presence of 40 g of ethanol liter−1. In this case, maximal activities were 3,900 and 4,660 nkat ml−1, respectively, corresponding to 1 and 1.2 g of laccase per liter and thus representing the highest laccase activities reported for recombinant fungal strains. These results suggest that P. cinnabarinus may be a host of choice for the production of other proteins as well. |
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