Purification and biochemical characterization of a transglucosilating beta-glucosidase of Stachybotrys strain.
The filamentous fungus Stachybotrys sp has been shown to possess a rich beta-glucosidase system composed of five beta-glucosidases. One of them was already purified to homogeneity and characterized. In this work, a second beta-glucosidase was purified and characterized. The filamentous fungal A19 strain was fed-batch cultivated on cellulose, and its extracellular cellulases (mainly beta-glucosidases) were analyzed. The purified enzyme is a monomeric protein of 78 kDa molecular weight and exhibits optimal activity at pH 6.0 and at 50 degrees C. The kinetic parameters, K (m) and V (max), on para-nitro-phenyl-beta-D: -glucopyranosid (p-NPG) as a substrate were, respectively, 1.846 +/- 0.11 mM and 211 +/- 0.08 micromol min(-1) ml(-1). One interesting feature of this enzyme is its high stability in a wide range of pH from 4 to 10. Besides its aryl beta-glucosidase activity towards salicin, methylumbellypheryl-beta-D: -glucoside (MU-Glc), and p-NPG, it showed a true beta-glucosidase activity because it splits cellobiose into two glucose monomers. This enzyme has the capacity to synthesize short oligosaccharides from cellobiose as the substrate concentration reaches 30% with a recovery of 40%. We give evidences for the involvement of a transglucosylation to synthesize cellotetraose by a sequential addition of glucose to cellotriose.
Saibi
Walid
W
Laboratoire de Génétique Moléculaire des Eucaryotes, Centre de biotechnologie de Sfax, CBS, route Sidi Mansour, Sfax, 3061, Tunisia.
Amouri
Bahia
B
Gargouri
Ali
A
D001709:Biotechnology; D044602:Cellulases / Q000378:metabolism; D002482:Cellulose / Q000031:analogs & derivatives* / Q000378:metabolism; D003470:Culture Media; D004795:Enzyme Stability; D005947:Glucose / Q000378:metabolism; D007700:Kinetics; D013191:Stachybotrys / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D013780:Tetroses / Q000378:metabolism*; D001617:beta-Glucosidase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism
D003470:Culture Media; D013780:Tetroses; C048468:cellotetraose; D002482:Cellulose; D044602:Cellulases; D001617:beta-Glucosidase; D005947:Glucose