黄孢原毛平革菌抗营养阻遏产漆酶诱变育种及其产酶特性

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

摘要:

摘要：【目的】筛选能抗营养阻遏产漆酶的黄孢原毛平革菌，论证其产漆酶的确定性及抗营养阻遏产木质素酶的可行性，为白腐菌产酶代谢调控、木质素降解机理的研究奠定基础。【方法】利用重复紫外诱变法，以愈创木酚富氮鉴别培养基筛选目标菌株；比较不同营养条件下菌体生长与产酶动力学差异研究产酶营养调控机理；通过热处理、排除锰离子和加入过氧化氢酶等不同措施论证黄孢原平毛平革菌能否产生漆酶。【结果】3种不同方法均证实选育到的pcR5305和pcR5324菌株在限氮与富氮条件下均能产生漆酶，pcR5305和pcR5324在限氮条件

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.