Trichoderma reesei

About: Trichoderma reesei is a research topic. Over the lifetime, 3832 publications have been published within this topic receiving 152877 citations. The topic is also known as: Trichoderma reesi.

Sequence of the cloned pyr4 gene of Trichoderma reesei and its use as a homologous selectable marker for transformation.

Abstract: We have cloned and sequenced the Trichoderma reesei pyr4 gene encoding orotidine-5′-monophosphate decarboxylase. Comparison of this sequence with that of the equivalent gene from other filamentous fungi suggests that T. reesei is closely related to Cephalosporium acremonium and Neurospora crassa. The cloned pyr4 gene has been used as a homologous selectable marker for transformation of T. reesei. The majority of transformants obtained with circular plasmid were mitotically unstable and contained non-integrated plasmid molecules, sometimes in addition to plasmid integrated in the genome, Linearization of plasmid prior to transformation decreased the transformation frequency but increased the proportion of stable transformation obtained.

Expression of Trichoderma reesei β-Mannanase in Tobacco Chloroplasts and Its Utilization in Lignocellulosic Woody Biomass Hydrolysis

Abstract: Lignocellulosic ethanol offers a promising alternative to conventional fossil fuels. One among the major limitations in the lignocellulosic biomass hydrolysis is unavailability of efficient and environmentally biomass degrading technologies. Plant-based production of these enzymes on large scale offers a cost-effective solution. Cellulases, hemicellulases including mannanases and other accessory enzymes are required for conversion of lignocellulosic biomass into fermentable sugars. β-mannanase catalyzes endo-hydrolysis of the mannan backbone, a major constituent of woody biomass. In this study, the man1 gene encoding β-mannanase was isolated from Trichoderma reesei and expressed via the chloroplast genome. PCR and Southern hybridization analysis confirmed site-specific transgene integration into the tobacco chloroplast genomes and homoplasmy. Transplastomic plants were fertile and set viable seeds. Germination of seeds in the selection medium showed inheritance of transgenes into the progeny without any Mendelian segregation. Expression of endo-β-mannanase for the first time in plants facilitated its characterization for use in enhanced lignocellulosic biomass hydrolysis. Gel diffusion assay for endo-β-mannanase showed the zone of clearance confirming functionality of chloroplast-derived mannanase. Endo-β-mannanase expression levels reached up to 25 units per gram of leaf (fresh weight). Chloroplast-derived mannanase had higher temperature stability (40°C to 70°C) and wider pH optima (pH 3.0 to 7.0) than E.coli enzyme extracts. Plant crude extracts showed 6–7 fold higher enzyme activity than E.coli extracts due to the formation of disulfide bonds in chloroplasts, thereby facilitating their direct utilization in enzyme cocktails without any purification. Chloroplast-derived mannanase when added to the enzyme cocktail containing a combination of different plant-derived enzymes yielded 20% more glucose equivalents from pinewood than the cocktail without mannanase. Our results demonstrate that chloroplast-derived mannanase is an important component of enzymatic cocktail for woody biomass hydrolysis and should provide a cost-effective solution for its diverse applications in the biofuel, paper, oil, pharmaceutical, coffee and detergent industries.

Metabolic engineering of inducer formation for cellulase and hemicellulase gene expression in Trichoderma reesei.

Abstract: The filamentous fungus T. reeseiis today a paradigm for the commercial scale production of different plant cell wall degrading enzymes mainly cellulases and hemicellulases. Its enzymes have a long history of safe use in industry and well established applications are found within the pulp, paper, food, feed or textile processing industries. However, when these enzymes are to be used for the saccharification of cellulosic plant biomass to simple sugars which can be further converted to biofuels or other biorefinery products, and thus compete with chemicals produced from fossil sources, additional efforts are needed to reduce costs and maximize yield and efficiency of the produced enzyme mixtures. One approach to this end is the use of genetic engineering to manipulate the biochemical and regulatory pathways that operate during enzyme production and control enzyme yield. This review aims at a description of the state of art in this area.

Exploring improved endoglucanase expression in Saccharomyces cerevisiae strains

Abstract: The endoglucanase I and II genes (egI or Cel7B and egII or Cel5A) of Trichoderma reesei QM6a were successfully cloned and expressed in Saccharomyces cerevisiae under the transcriptional control of the yeast ENO1 promoter and terminator sequences. Random mutagenesis of the egI-bearing plasmid resulted in a twofold increase in extracellular EGI activity. Both endoglucanase genes were co-expressed with the synthetic, codon-optimised cellobiohydrolase gene (s-cbhI) from T. reesei as well as the β-glucosidase gene (bgl1) from Saccharomycopsis fibuligera in S. cerevisiae. Extracellular endoglucanase activity was lower when co-expressed with s-cbhI or bgl1. Recombinant strains were able to hydrolyse phosphoric acid swollen cellulose, generating mainly cellotriose, cellobiose and glucose. Cellobiose accumulated, suggesting the β-glucosidase activity to be the rate-limiting factor. As a consequence, the recombinant strains were unable to produce enough glucose for growth on amorphous cellulose. The results of this study provide insight into further optimisation of recombinantly expressed cellulase combinations for saccharification and fermentation of cellulose to ethanol.