The Enzyme System Reforming For High Efficiency Degradation Lignocellulose

Because of Trichoderma reesei unusual protein yield ability,i is considered to be the best host for homologous or heterologous protein production.The production of heterologous proteins in Trichoderma reesei is lower than homologous protein,therefore,to explain whether exogenous protein production depends on exogenous gene promoter is very important.The expression of Trichoderma reesei cellulase is high,but the production of?-glucosidase is very low.The purpose is to explore the Trichoderma reesei promoter for exogenous gene and construct efficient lignocellulose degradation strains.Trichoderma reesei cbh?,xyn?,cbhl?,xyn?,xyn? gene are replaced by Aspergillus niger ?-glucosidase gene by homologous recombination,and then we measured enzyme activity and saccharification efficiency between wild strain and mutant strains.The data shows that mutant strains using the homologous replacement of Trichoderma reesei is better.The promoter from endogenous protein to initiate exogenous gene can get better results;the comprehensive data including the extracell-ular protein,cellulase activity and other experimental data show that,the promoters from Trichoderma reesei xyn?,xyn? gene whose expression quantity is not high,ho-wever,the production of exogenous genes of ?-glucosidase is very high.We obtained Trichoderma reesei who has complete enzyme system achieved high saccharification efficiency.We explored the component of cellulase to find out cellulase degradation mecha-nism.We constructed Trichoderma reesei hetergolous expressing ?-glucosidase from Aspergillus aculeatus by homologous recombination.The successful realization of the heterologous expression(3-glucosidase,the production of ?-glucosidase and Sacchar-ification efficiency of extracellular protein was greatly enhanced.There is a big difference between cellulase degradation efficiency between Tri-choderma strain QM6a and T1.We measured and found that enzyme activity ratio is not consistent,but the effect of degradation efficiency is whether effected by other re-asons.So we analyzed the clustering of extracellular proteins in two Trichoderma re-esei strains using blue-native gel electrophoresis.We analyzed the saccharification ef-ficiency and the results of polymer depolymerization experiments,to further explain the degradation mechanism of cellulase.By blue native gel(BN-PAGE)technology a-nd mass spectrometry analysis,we found that cellulase polymerultimers are mainly composed of cellulose,hemicellulase,cellulase different modified forms.Specific reason still needs further exploration;We found the saccharification efficiency is different significantly.Especially the saccharification efficiency of T1 is much higher than that of QM6a through depolymerization experiments,further proofing of the existence of cellulosemultimeric forms of aggregation,and the important role of degradation of lingocellulose.We explored the cellulase degradation mechanism by analyzed cellulose multi-meric composition.From the perspective of cellulase multimer formation,the explor-ation of the mechanism of degradation enzyme of Trichoderma reesei strain and the mutant strains by using BN-PAGE technology lay a very important theoretical found-ation for further improving the cellulose.