Constitutive expression of the Trichoderma reesei beta-1,4-xylanase gene (xyn2) and the beta-1,4-endoglucanase gene (egI) in Aspergillus niger in molasses and defined glucose media

• Published in: Applied microbiology and biotechnology Vol. 58; no. 4; pp. 461 - 468
• Main Authors: Rose, S.H, Van Zyl, W.H
• Format: Journal Article
• Language: English
• Published: Berlin Springer 01.03.2002
• Subjects: Aspergillus awamori; Aspergillus niger; Aspergillus niger - genetics; beta-Galactosidase - genetics; beta-Galactosidase - metabolism; Biological and medical sciences; Biotechnology; Cloning, Molecular; Culture Media; Electrophoresis, Polyacrylamide Gel; Endo-1,4-beta Xylanases; endo-1,4-beta-glucanase; Enzyme engineering; Fermentation; Fundamental and applied biological sciences. Psychology; Gene Expression; genes; glucose; Glucose - metabolism; Glycoside Hydrolases; Hydrogen-Ion Concentration; Methods. Procedures. Technologies; Molasses; Plasmids - genetics; Production of selected enzymes; proteins; transcription (genetics); Trichoderma - enzymology; Trichoderma - genetics; Trichoderma reesei; xylanases; Xylosidases - genetics; Xylosidases - metabolism; Recombinant microorganism; Molasses; Enzyme; Endo-1,4-β-xylanase; Gene expression; Trichoderma reesei; Fungi; Enzymatic activity; Aspergillus niger; Cellulase; Glycosidases; Production; Microorganism culture; Hydrolases; Fungi Imperfecti; O-Glycosidases; Physicochemical properties; Thallophyta
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s00253-001-0922-3

The xylanase II (xyn2)- and endoglucanase I (egI)-encoding regions of Trichoderma reesei QM6a were successfully expressed in Aspergillus niger D15 under the transcriptional control of the glyceraldehyde-6-phosphate dehydrogenase (gpd) promoter from A. niger and the glaA terminator of Aspergillus awamori. A stable xyn2 transformant produced beta-xylanase activity of 8,000 nkat/ml and 5,000 nkat/ml in shake-flask cultures containing defined or 20% (v/v) molasses medium, respectively. The recombinant Xyn2 enzyme expressed highest activity at pH 5-6 and 50-60 degrees C and retained more than 75% of its activity after 3 h of incubation at 50 degrees C. A stable egI transformant produced endo-beta-1,4-glucanase activity of 2,300 nkat/ml in shake-flask cultures containing defined media and about half the activity in 20% molasses medium. Maximum endoglucanase activity was obtained at pH 5 and 60 degrees C. Both Xyn2 and EgI retained >80% activity after incubation at 50 degrees C for 3 h. The heterologous Xyn2 and EgI represent a significant portion of the total extracellular proteins produced.