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.

Stability of the Cellulase of Trichoderma reesei under use conditions

Abstract: Enzyme stability studies have been reinvestigated under the conditions used for cellulose hydrolysis (pH 48, 50 degrees C, 24 hr) The cellobiohydrolase (CBH) component as measured on Avicel is less stable than other enzymes of the cellulase complex, and is 60% inactivated by merthiolate (and other Hg compounds) under the above conditions Endo-beta-1,4-glucanase is much more stable, and more resistant to merthiolate and other compounds Under unshaken conditions the Avicelase of the Rutgers strain C 30 shows greater stability to heat than that of other available strains Biocides must be selected not only for their ability to prevent contamination, but also for their compatibility with cellulases Tetracycline and chlortetracycline are inexpensive, effective in very low concentrations, have no harmful effect on the enzymes, and are compatible with the yeasts that subsequently grow on the sugar solutions to produce alcohol Attempts have been made to stabilize the enzymes by chemical modification in such a way as to maintain their solubility Glutaraldehyde treatment greatly increased the enzyme size, lowered the pI values, and gave a slight shift in the pH activity curve There was, unfortunately, no increase in enzyme stability, and the activity of enzymes on solid celluloses was adversely affected Shaking greatly reduced the hydrolysis of Avicel by Trichoderma reesei C 30 enzyme The adverse effect was accompanied by a decrease in recoverable enzyme and protein

Degradation of Xylan to D-Xylose by Recombinant Saccharomyces cerevisiae Coexpressing the Aspergillus niger β-Xylosidase (xlnD) and the Trichoderma reesei Xylanase II (xyn2) Genes

Abstract: Plant cell walls, the major reservoir of fixed carbon in nature, contain three major polymers: cellulose (insoluble fibers of β-1,4-glucan), hemicellulose (noncellulosic polysaccharides including xylans, mannans, and glucans) and lignin (a complex polyphenolic structure) (1, 45). β-1,4-Xylans are found mainly in secondary walls of plants and can represent up to 35% of the total dry weight in certain plants. Xylan is a complex polysaccharide consisting of a backbone of β-d-1,4-linked xylopyranoside units substituted with acetyl, glucuronosyl, and arabinosyl side chains. Endo-β-xylanases (EC 3.2.1.8) act on xylans and xylo-oligosaccharides, producing mainly mixtures of xylooligosaccharides (4, 23). β-d-Xylosidases (EC 3.2.1.37) hydrolyze xylooligosaccharides, produced through the action of β-xylanases, to d-xylose. Many bacterial and fungal species are able to utilize xylans as a carbon source (18). Strains of the fungi Trichoderma and Aspergillus secrete large amounts of efficient xylan-degrading enzymes (8, 16, 51). Recently, interest in β-xylanases has increased because of their application in biobleaching (30, 44) and the food (31) and animal feed (3, 34, 47) industry. Trichoderma reesei is a filamentous mesophilic fungus that is well known for its cellulolytic and xylanolytic enzymatic activities (12, 43). The two major inducible endo-β-xylanases secreted by this fungus are Xyn1 and Xyn2 (46). They are both relatively small protein molecules, with molecular masses of 19 and 21 kDa, respectively, but Xyn2 represents more than 50% of the total xylanolytic activity of T. reesei cultivated on xylan. Fungi of the genus Aspergillus are also efficient producers of cellulose- and xylan-degrading enzymes, regulated at the transcriptional level by the XlnR activator (49). The two endo-β-xylanases and the β-xylosidase in A. niger are encoded by xlnB, xlnC, and xlnD, respectively. The xlnD gene contains an open reading frame of 2,412 nucleotides, which encodes a protein of 804 amino acids with a predicted molecular mass of 85 kDa. The protein is N glycosylated and contains 15 potential N-glycosylation sites (48). Sequence similarity was found to β-glycosidases (β-xylosidase and β-glucosidases) of family 3, which include enzymes from both bacterial and fungal origins (20, 33, 35, 48). The condensation reaction of this β-xylosidase has been used for the synthesis of disaccharides such as β,β-1,1-xylodisaccharide, β-1,4-xylodisaccharide (xylobiose), β-1,2-xylodisaccharide, α-1,4-xylodisaccharide, and β-1,3-xylodisaccharide (17). S. cerevisiae has been successfully used for the production of related fungal β-xylosidase and β-glucosidases belonging to family 3 (7, 32, 33). Different Candida species (C. maltosa, C. tropicalis, and C. utilis) are currently used in industry for the production of single-cell protein and ethanol from steamed hemicellulose (21). Even though these Candida strains are able to ferment d-xylose, none of them are able to tolerate the same levels of ethanol as Saccharomyces cerevisiae does, and, furthermore, they cannot ferment hexose sugars as effectively. However, the main disadvantage of S. cerevisiae is the fact that it cannot hydrolyze xylan or utilize or ferment d-xylose, the main component of xylan. While research is continuing on the development of a S. cerevisiae strain able to ferment d-xylose (9, 14, 28, 50), we are working toward the construction of strains able to break down the xylan backbone to its monomeric constituent, d-xylose. In this paper, we describe the molecular cloning of the A. niger xlnD gene and its expression in S. cerevisiae. Expression and coexpression of xlnD and xyn2 from T. reesei in yeast was obtained with the aid of multicopy plasmids using the derepressible S. cerevisiae alcohol dehydrogenase II gene promoter (ADH2P) and terminator (ADH2T) sequences (38). The enhanced production of both the recombinant enzymes in non-selective complex medium, without the risk of losing the episomal vector, was obtained by constructing autoselective recombinant fur1 S. cerevisiae strains (29).

Differential Expression of the Vegetative and Spore‐Bound Hydrophobins of Trichoderma Reesei Cloning and Characterization of the Hfb2 Gene

Abstract: The hfb2 gene encoding the hydrophobin HFBII of the filamentous fungus Trichoderma reesei was isolated by heterologous hybridization using the vegetative hydrophobin I, hfb1, gene of T. reesei as a probe. The hfb2 gene codes for a typical fungal secreted hydrophobin of 71 amino acids containing eight cysteine residues. The amino acid similarity towards HFBI is 69%. The HFBII protein was isolated from the fungal spores by extraction with trifluoroacetic acid/acetonitrile solution, and by bubbling from the lactose-based culture medium. Expression of the hfb1 and hfb2 genes is divergent. hfb1 expression was only observed in vegetative cultures on glucose-containing and sorbitol-containing media. It was not expressed on media containing complex plant polysaccharides, cellulose, xylan, cellobiose or lactose, whereas hfb2 was highly expressed in vegetative cultures on these media. Expression of hfb2 was also strongly induced by N and C starvation, by light and in conidiating cultures.

Three-dimensional structure of a thermostable native cellobiohydrolase, CBH IB, and molecular characterization of the cel7 gene from the filamentous fungus, Talaromyces emersonii.

Abstract: The X-ray structure of native cellobiohydrolase IB (CBH IB) from the filamentous fungus Talaromyces emersonii, PDB 1Q9H, was solved to 2.4 A by molecular replacement. 1Q9H is a glycoprotein that consists of a large, single domain with dimensions of ≈ 60 A × 40 A × 50 A and an overall β-sandwich structure, the characteristic fold of Family 7 glycosyl hydrolases (GH7). It is the first structure of a native glycoprotein and cellulase from this thermophilic eukaryote. The long cellulose-binding tunnel seen in GH7 Cel7A from Trichoderma reesei is conserved in 1Q9H, as are the catalytic residues. As a result of deletions and other changes in loop regions, the binding and catalytic properties of T. emersonii 1Q9H are different. The gene (cel7) encoding CBH IB was isolated from T. emersonii and expressed heterologously with an N-terminal polyHis-tag, in Escherichia coli. The deduced amino acid sequence of cel7 is homologous to fungal cellobiohydrolases in GH7. The recombinant cellobiohydrolase was virtually inactive against methylumberiferyl-cellobioside and chloronitrophenyl-lactoside, but partial activity could be restored after refolding of the urea-denatured enzyme. Profiles of cel7 expression in T. emersonii, investigated by Northern blot analysis, revealed that expression is regulated at the transcriptional level. Putative regulatory element consensus sequences for cellulase transcription factors have been identified in the upstream region of the cel7 genomic sequence.