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KMID : 0545120080180050933
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Journal of Microbiology and Biotechnology
2008 Volume.18 No. 5 p.933 ~ p.941
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Purification and Characterization of an Extracellular β-Glucosidase from Monascus purpureus
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Daroit Daniel J.
Aline Simonetti
Plinho F. Hertz
Adriano Brandelli
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Abstract
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An extracellular β-glucosidase produced by Monascus purpureus NRRL1992 in submerged cultivation was purified by acetone precipitation, gel filtration, and hydrophobic interaction chromatography, resulting in a purification factor of 92-fold. A 22 central-composite design (CCD) was performed to find the best temperature and pH conditions for enzyme activity. Maximum activity was observed in a wide range of temperature and pH values, with optimal conditions set at 50oC and pH 5.5. The β-glucosidase showed moderate thermostability, was inhibited by HgCl2, K2CrO4, and K2Cr2O7, whereas other reagents including β- mercaptoethanol, SDS, and EDTA showed no effect. Activity was slightly stimulated by low concentrations of ethanol and methanol. Hydrolysis of p-nitrophenyl-β-D-glucopyranoside (pNPG), cellobiose, salicin, n-octyl-β-D-glucopyranoside, and maltose indicates that the β-glucosidase has broad substrate specificity. Apparently, glucosyl residues were removed from the nonreducing end of p-nitrophenyl-β-Dcellobiose. β-Glucosidase affinity and hydrolytic efficiency were higher for pNPG, followed by maltose and cellobiose. Glucose and cellobiose competitively inhibited pNPG hydrolysis.
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KEYWORD
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β-Glucosidase, Monascus purpureus, characterization, thermostability, substrate specificity, kinetic constants
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