Breeding and characterization of laccase-producing Phanerochaete chrysosporium mutant resistant to nutritional repression

Abstract:

Abstract: ［Objective］To screen Phanerochaete chrysosporium mutants resisting nutritional repression and to characterize laccase produced by the mutants. [Methods] We used repeated UV mutagenesis and screened the mutant strains by using the guaiacol nitrogen sufficient differential medium. We characterized enzymes production mechanism of the nutritional regulation through comparing the differences of cell growth and enzyme-production kinetics under different nutritional conditions；We validated production of laccase by Phanerochaete chrysosporium through measurements of the heat treatment, removal of manganese ion and addition of the catalase. ［Results］Three different methods were validated that both strains of pcR5305 and pcR5324 can produce laccase under the nitrogen limitation (N-L) and nitrogen sufficient (N-S) conditions. Under the N-L conditions, pcR5305 can produce 203.5 U/L laccase and pcR5324 can produce 187.6 U/L laccase；Under the N-S conditions，pcR5305 can produce 220.6 U/L laccase and pcR5324 can produce 183.9 U/L laccase. The original strain pc530 only can produce very little laccase under either conditions. The laccase-production regulation mechanisms of the two strains are different: Production of laccase and the cell growth by pcR5305 are in synchronism. However production of the laccase by pcR5324 is repressed by nutrition. Both strains have the capacity of resisting nutritional repression and produce lignin peroxidase and manganese peroxidase with high yield. (LiP 1343.2、MnP 252.2 U/L and LiP 1169.5、MnP 172.4 U/L respectively). [Conclusion] The mutants of Phanerochaete chrysosporium can produce laccase. At same time they showed the capacity of resisting nutritional repression and production of laccase, lignin peroxidase and manganese peroxidase. Our results posses high value for production, application and fundamental research. We provided new strains and established a very good foundation for the further research of metabolic regulation of ligninolytic enzymes production.