Purification and characterization of an endo-1,3-beta-glucanase from Aspergillus fumigatus.
An endo-1,3-beta-glucanase was purified from a cell wall autolysate of Aspergillus fumigatus. This beta-glucanase activity was associated with a glycosylated 74-kDa protein. Using a sensitive colorimetric assay and a high-performance anion-exchange chromatography with a pulsed electrochemical detector for product analysis, it was shown that the endoglucanase hydrolysed exclusively linear 1,3-beta-glucan chains, had an optimum pH of 7.0 and an optimum temperature of 60 degrees C. A substrate kinetic study gave a Km value of 0.3 mg/ml for soluble (laminarin and laminari-oligosaccharides) and 1.18 mg/ml for insoluble (curdlan) 1,3-beta-glucan. Laminari-oligosaccharide degradation, analysed by HPLC, showed that the endoglucanase bind to the subtrate at several positions and suggested that the active site of the enzyme recognized five glucose units linked by a 1,3-beta bond. The association of the present endo-1,3-beta-glucanase with the cell wall of A. fumigatus suggests a putative role for this enzyme during cell-wall morphogenesis.
Fontaine
T
T
Institut Pasteur, Laboratoire des Aspergillus, Paris, France.
Hartland
R P
RP
Beauvais
A
A
Diaquin
M
M
Latge
J P
JP
D001232:Aspergillus fumigatus / Q000201:enzymology*; D002473:Cell Wall / Q000201:enzymology; D005656:Fungal Proteins / Q000302:isolation & purification; D004693:Glucan Endo-1,3-beta-D-Glucosidase / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D005936:Glucans / Q000378:metabolism; D006031:Glycosylation; D006863:Hydrogen-Ion Concentration; D008562:Membrane Glycoproteins / Q000302:isolation & purification; D008970:Molecular Weight; D011134:Polysaccharides / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature; D047071:beta-Glucans
D005656:Fungal Proteins; D005936:Glucans; D008562:Membrane Glycoproteins; D011134:Polysaccharides; D047071:beta-Glucans; C025363:mannoproteins; C038459:curdlan; C008247:laminaran; D004693:Glucan Endo-1,3-beta-D-Glucosidase