A thermostable β-glucosidase. The enzyme catalyses the transfer of D-glucose residues in disaccharides, alkyl glycosides or other aglycone-linked glycosides to water (hydrolysis) or to alcohols depending on reaction conditions.
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Activity determination: Glycosidase activity was determined at 85°C in a 20 mM citrate phosphate buffer at pH 5.6 and incubated for 5 minutes with 2.92 mM final concentration of p-nitrophenyl-β-D-glucanopyranoside.
Unit definition: One unit (U) of enzyme activity is the amount that leads to the release of 1 μmol of p-nitrophenyl from p-nitrophenyl-β-D-glucanopyranoside (pNPG) per minute.
Synonyms: β-glucosidase, glycoside hydrolase, β-D-glucoside glucohydrolase
Protein family: Glycosyl hydrolase family 3 ( GH3 ) – CAZy database GH3 family
Enzyme classification: EC 126.96.36.199
Source: The thermophilic bacteria Thermotoga neopolitana
Protein sequence: NBCI protein entry
Structural information: The crystal structure of β-glucosidase from Thermotoga neopolitana has been determined to 2.05 Å resolution (Pozzo et al. 2010). – PDB entry 2X41
Figure 1: Crystal structure of β-glucosidase 3B from Thermotoga neopolitana
Substrates and Applications:
Polyphenol glycoside hydrolysis: The enzyme may be used for hydrolysis of various glycosides such as polyphenol glycosides including naturally occurring antioxidants such as quercetin-glycosides found in various vegetables (Fig. 2). For complete hydrolysis of 1 μmol of quercetin-4-glycoside in 5 minutes at 80°C and pH 5.5, about 28 pmol (~25 μg) enzyme was needed (Turner et al 2006)
Alkyl glycoside synthesis: The enzyme may be used for transglycolysation with alcohols as acceptors such as in synthesis of alkyl glycosides. At 60°C, the ratio of alcoholysis to hydrolysis is 5.1 with a pNPG conversion rate of 153U/mg whereas at 90°C the ratio of alcoholysis to hydrolysis is 4.5 and the pNPG conversion rate of is 831U/mg. (Turner et al. 2007)
Properties of ThermoActive™ β-glucosidase:
Figure 3. Example of substrate specificity: quercetin
Turner C, Turner P, Jacobson G, Waldebäck M, Sjöberg P, Nordberg Karlsson E & Markides K. (2006). Subcritical water extraction and β-glucosidase catalyzed hydrolysis of quercetin in onion waste. Green Chem. 8: 949–959.
Turner P, Svensson D, Adlercreutz P & Karlsson EN (2007). A novel variant of Thermotoga neapolitana β-glucosidase B is an efficient catalyst for the synthesis of alkyl glucosides by transglycosylation. J. Biotechnol.130: 67–74.