Highly Efficient Production of Laccase by the Basidiomycete Pycnoporus cinnabarinus

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.

Enhanced benzaldehyde formation by a monokaryotic strain of Pycnoporus cinnabarinus using a selective solid adsorbent in the culture medium

A monokaryotic strain of the white-rot fungus Pycnoporus cinnabarinus was shown to produce, in a 2-L bioreactor culture, 100 mg·L-1 benzaldehyde (bitter almond aroma) from L-phenylalanine with a productivity of 33 mg·L-1·day-1. The addition of HP20 resin, a styrene divinylbenzene copolymer highly selective for benzaldehyde, enabled an eightfold increase in the production of benzaldehyde and a twofold increase in productivity. In the presence of HP20 resin, the production of 790 mg·L-1 benzaldehyde was concomitant with the synthesis of cinnamic acid derivatives of high organoleptic notes such as cinnamaldehyde, cinnamyl alcohol, and methyl cinnamate.Key words : benzaldehyde, L-phenylalanine, Pycnoporus cinnabarinus, adsorbents.

Carbon source dependent differences in the composition of the cell walls of the basidiomycete Picnoporus cinnabarinus

A wild-type monokaryotic strain of Picnoporus cinnabarinus grown on glucose produced shorter and thicker hyphae than cultures grown on acetate. Colonies from glucose media were smaller and more compact than acetate-grown colonies. Chemical and enzymatic analysis of the isolated cell wall of both morphological types showed that the amount of amino sugars and the ratio glucosamine:galactosamine were higher in the acetate-grown cells. This may be the cause of morphological differences observed.