Molecular Characterization of a Highly-Active Thermophilic β-Glucosidase from Neosartorya fischeri P1 and Its Application in the Hydrolysis of Soybean Isoflavone Glycosides

Abstract

Isoflavone occurs abundantly in leguminous seeds in the form of glycoside and aglycone. However, isoflavone glycoside has anti-nutritional effect and only the free type is beneficial to human health. In the present study we identified a β-glucosidase from thermophilic Neosartorya fischeri P1, termed NfBGL1, capable of efficiently converting isoflavone glycosides into free isoflavones. The gene, belonging to glycoside hydrolase family 3, was successfully overexpressed in Pichia pastoris at high cell density in a 3.7-l fermentor. Purified recombinant NfBGL1 had higher specific activity (2189±1.7 U/mg) and temperature optimum (80°C) than other fungal counterparts when using p-nitrophenyl β-d-glucopyranoside as the substrate. It retained stable at temperatures up to 70°C and over a broad pH range of 3.0−10.0. NfBGL1 had broad substrate specificity including glucosidase, cellobiase, xylanase and glucanase activities, and displayed preference for hydrolysis of β-1,2 glycosidic bond rather than β-1,3, β-1,4, β-1,6 bonds. The enzyme showed high bioconversion ability for major soybean isoflavone glycosides (daidin, gensitin and glycitin) into free forms. These properties make NfBGL1 potential for the wide use in the food, feed, pharmacy and biofuel industries.

Microbial β-Glucosidases: Screening, Characterization, Cloning and Applications

“…Microbial β-glucosidases from different sources have been characterized from different sources for molecular properties [155].…”

Biochemical characterization of three distinct polygalacturonases from Neosartorya fischeri P1

“…Neosartorya fischeri strain P1 is an excellent carbohydrate-active enzyme (CAZyme) producer Yang et al, 2014). It can utilize pectin as the sole carbon source and secret extracellular pectinases.…”

Alkali-tolerant β-glucosidase produced by newly isolated Aspergillus fumigatus WL002 from rotten wood

“…Many b-glucosidase-producing organisms have been isolated from different sources (Bajaj et al, 2009;Kaur et al, 2007;Matkar et al, 2013;Saha and Bothast, 1996;Yang et al, 2014). However, the optimum time for maximum enzyme production by different microbes was relatively variable.…”

“…The enzymatic activities against pNPG (Sigma) and oNPG (Sigma) were evaluated, as described by Yang et al (2014). In particular, the reactions were conducted in 50 mM glycine-NaOH buffer (pH 9.0) at 50 C for 10 min with 5 mM substrate and then terminated by supplementing 1.0 ml of 1 M Na 2 CO 3 .…”

“…In particular, the DNS method described by Miller (1959) was conducted at 50 C and pH 9.0 to evaluate the specificity to salicin, esculin, Avicel, CMC-Na, and PASC; by contrast, the specificity to pNPG and oNPG was assayed at 50 C and pH 9.0 according to the method described by Yang et al (2014).…”