Domingues, Lucília

Ilmén, M.

Penttilä, M.

Ribeiro, Orquídea

Wiebe, M.

Universidade do Minho: RepositoriUM

Expression of Trichoderma reesei cellulases in the fungus Ashbya gossypii

Topic: Filamentous and Non-filamentous Fungi in Biotechnology and DiseaseTitle: EXPRESSION OF TRICHODERMA REESEI CELLULASES IN THEFUNGUS ASHBYA GOSSYPIIAuthor(s): O. Ribeiro1, M. Wiebe2, M. Ilmén2, L. Domingues1, M. Penttilä2Institute(s): 1IBB - Institute for Biotechnology and Bioengineering - Centre of BiologicalEngineering, University of Minho, Braga, Portugal, 2VTT TechnicalResearch Centre of Finland, Espoo, FinlandText: Background: Due to its natural ability to produce riboflavin (vitamin B2),Ashbya gossypii is a widely used microorganism in industry. Additionally, itpresents several other interesting features, such as, the smallest knowneukaryotic genome, lack of extensive duplication of chromosomalsegments and straight forward molecular tools for genetic manipulation.Together, these make it an interesting microorganism, not only forriboflavin production, but also for other applications such as host forrecombinant protein production.The enzymatic degradation of cellulose is of great importance to thecarbon cycle of the biosphere, since is the most abundant biopolymer.Bioconversion of cellulose to soluble sugars is catalyzed by a group ofenzymes called cellulases.Objectives:1. Evaluate the potential of A. gossypii as a host forrecombinant protein production, in particular, cellulases CBHI and EGIfrom Trichoderma reesei. 2. Partially characterize the heterologouscellulases secreted by A. gossypiiMethods:1. Expression plasmids for production of T. reesei cellulases,namely, CBHI and EGI were used to transform A. gossypii. 2. Activity onsoluble substrates was determined. 3. The secreted cellulases werepartially characterized.Results: Recombinant EGI was produced by A. gossypii in complex, butnot in defined medium. Production was similar at 30 and 24°C. Similaramounts of enzyme activity, were produced by A. gossypii and S.cerevisiae. CBHI activity was not detectable, but the protein could bedetected by Western blotting. The cellulases secreted by A. gossypii had adifferent glycosylation pattern than the same proteins secreted by S.cerevisiae.Conclusions: These initial results demonstrate that CBHI and EGI aresecreted by recombinant A. gossypii into the extracellular medium withsimilar titers to those secreted by S. cerevisiae. Further optimization of thegenetic constructions and culture conditions could lead to improvedsecretion.PreferredPresentation Type: Poster Presentation

http://repositorium.sdum.uminho.pt/bitstream/1822/10370/1/Ribeiro_FEMS2009.pdf

Expression of Trichoderma reesei cellulases in the fungus Ashbya gossypii

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Universidade do Minho: RepositoriUM

Universidade do Minho: RepositoriUM