Constitutive expression of the Trichoderma reesei beta-1,4-xylanase gene (xyn2) and the beta-1,4-endoglucanase gene (egl) in Aspergillus niger in molasses and defined glucose media.
The xylanase II (xyn2)- and endoglucanase I (egl)-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 egl transformant produced endo-P-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.
Rose
S H
SH
Department of Microbiology, University of Stellenbosch, South Africa.
van Zyl
W H
WH
D001234:Aspergillus niger / Q000235:genetics; D003001:Cloning, Molecular; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D043364:Endo-1,4-beta Xylanases; D015870:Gene Expression; D005947:Glucose / Q000378:metabolism*; D006026:Glycoside Hydrolases; D006863:Hydrogen-Ion Concentration; D008965:Molasses; D010957:Plasmids / Q000235:genetics; D014242:Trichoderma / Q000201:enzymology* / Q000235:genetics; D014995:Xylosidases / Q000235:genetics / Q000378:metabolism*; D001616:beta-Galactosidase / Q000235:genetics* / Q000378:metabolism
D003470:Culture Media; D006026:Glycoside Hydrolases; D014995:Xylosidases; D001616:beta-Galactosidase; D043364:Endo-1,4-beta Xylanases; C094192:arabinogalactan endo-1,4-beta-galactosidase; D005947:Glucose