Heterologous expression of Trametes versicolor laccase in Pichia pastoris and Aspergillus niger |
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Authors: | Christina Bohlin Leif J Jönsson Robyn Roth Willem H van Zyl |
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Institution: | (1) Biochemistry, Division for Chemistry, Karlstad University, SE-651 88 Karlstad, Sweden;(2) Bio/Chemtek, CSIR, Private Bag X2, 1645 Modderfontein, South Africa;(3) Department of Microbiology, University of Stellenbosch, Private Bag X1, 7602 Matieland, South Africa |
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Abstract: | Convenient expression systems for efficient heterologous production of different laccases are needed for their characterization
and application. The laccase cDNAs lcc1 and lcc2 from Trametes versicolor were expressed in Pichia pastoris and Aspergillus niger under control of their respective glyceraldehyde-3-phosphate dehydrogenase promoters and with the native secretion signal
directing catalytically active laccase to the medium. P. pastoris batch cultures in shake-flasks gave higher volumetric activity (1.3 U/L) and a better activity to biomass ratio with glucose
than with glycerol or maltose as carbon source. Preliminary experiments with fed-batch cultures of P. pastoris in bioreactors yielded higher activity (2.8 U/L) than the shake-flask experiments, although the levels remained moderate
and useful primarily for screening purposes. With A. niger, high levels of laccase (2700 U/L) were produced using a minimal medium containing sucrose and yeast extract. Recombinant
laccase from A. nigher harboring the lcc2 cDNA was purified to homogeneity and it was found to be a 70-kDa homogeneous enzyme with biochemical and catalytic properties
similar to those of native T. versicolor laccase A. |
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Keywords: | Laccase heterologous expression Pichia pastoris Aspergillus niger |
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