Daroit Daniel J, Simonetti Aline, Hertz Plinho F, Brandelli Adriano
Laboratorio de Bioquimica e Microbiologia Aplicada, Departamento de Ciencia de Alimentos (ICTA), Universidade Federal do Rio Grande do Sul, 91501-970, Porto Alegre, RS, Brazil.
J Microbiol Biotechnol. 2008 May;18(5):933-41.
An extracellular beta-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 50 degrees and pH 5.5. The beta-glucosidase showed moderate thermostability, was inhibited by HgCl2, K2CrO4, and K2Cr2O7, whereas other reagents including beta- mercaptoethanol, SDS, and EDTA showed no effect. Activity was slightly stimulated by low concentrations of ethanol and methanol. Hydrolysis of p-nitrophenyl-beta-D-glucopyranoside (pNPG), cellobiose, salicin, n-octyl-beta-D-glucopyranoside, and maltose indicates that the beta-glucosidase has broad substrate specificity. Apparently, glucosyl residues were removed from the nonreducing end of p-nitrophenyl-beta-Dcellobiose. beta-Glucosidase affinity and hydrolytic efficiency were higher for pNPG, followed by maltose and cellobiose. Glucose and cellobiose competitively inhibited pNPG hydrolysis.
紫红曲霉NRRL1992在深层培养中产生的一种胞外β-葡萄糖苷酶，通过丙酮沉淀、凝胶过滤和疏水相互作用色谱法进行纯化，纯化倍数为92倍。采用22中心复合设计（CCD）来寻找酶活性的最佳温度和pH条件。在较宽的温度和pH值范围内观察到最大活性，最佳条件设定为50℃和pH 5.5。该β-葡萄糖苷酶表现出中等的热稳定性，受到HgCl2、K2CrO4和K2Cr2O7的抑制，而其他试剂包括β-巯基乙醇、SDS和EDTA则无影响。低浓度的乙醇和甲醇对活性有轻微的促进作用。对硝基苯基-β-D-吡喃葡萄糖苷（pNPG）、纤维二糖、水杨苷、正辛基-β-D-吡喃葡萄糖苷和麦芽糖的水解表明该β-葡萄糖苷酶具有广泛的底物特异性。显然，葡萄糖基残基从对硝基苯基-β-D-纤维二糖的非还原端被去除。β-葡萄糖苷酶对pNPG的亲和力和水解效率更高，其次是麦芽糖和纤维二糖。葡萄糖和纤维二糖竞争性抑制pNPG的水解。
