A Novel Thermostable GH3<i>β</i>-Glucosidase from<i>Talaromyce leycettanus</i>with Broad Substrate Specificity and Significant Soybean Isoflavone Glycosides-Hydrolyzing Capability

Li, Xinxin; Xia, Wei; Bai, Yingguo; Ma, Rui; Yang, Hong; Luo, Huiying; Shi, Pengjun

doi:10.1155/2018/4794690

Cited by 18 publications

(25 citation statements)

References 31 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…To the best of our knowledge, this value is only surpassed by those found for the recombinant PtBglu3 from Paecilomyces thermophila [22], and bgl3A, from Talaromyces leycettanus [23]. In general, most of the recombinant strains producing BGLs reach an activity between 1 and 50 U/mL, which highlights the expression levels of BGL-1 [5,14,15,[24][25][26][27]. It is important to emphasize that the activity determined for this recombinant BGL-1 was 35-fold higher than the total β-glucosidase activity detected in cultures of T. amestolkiae [14], which corresponds to the sum of the activities of BGL-1, BGL-2 and BGL-3.…”

Section: Discussionmentioning

confidence: 79%

A glucotolerant β-glucosidase from the fungus Talaromyces amestolkiae and its conversion into a glycosynthase for glycosylation of phenolic compounds

et al. 2020

View full text Add to dashboard Cite

Background The interest for finding novel β-glucosidases that can improve the yields to produce second-generation (2G) biofuels is still very high. One of the most desired features for these enzymes is glucose tolerance, which enables their optimal activity under high-glucose concentrations. Besides, there is an additional focus of attention on finding novel enzymatic alternatives for glycoside synthesis, for which a mutated version of glycosidases, named glycosynthases, has gained much interest in recent years. Results In this work, a glucotolerant β-glucosidase (BGL-1) from the ascomycete fungus Talaromyces amestolkiae has been heterologously expressed in Pichia pastoris, purified, and characterized. The enzyme showed good efficiency on p-nitrophenyl glucopyranoside (pNPG) (Km= 3.36 ± 0.7 mM, kcat= 898.31 s−1), but its activity on cellooligosaccharides, the natural substrates of these enzymes, was much lower, which could limit its exploitation in lignocellulose degradation applications. Interestingly, when examining the substrate specificity of BGL-1, it showed to be more active on sophorose, the β-1,2 disaccharide of glucose, than on cellobiose. Besides, the transglycosylation profile of BGL-1 was examined, and, for expanding its synthetic capacities, it was converted into a glycosynthase. The mutant enzyme, named BGL-1-E521G, was able to use α-d-glucosyl-fluoride as donor in glycosylation reactions, and synthesized glucosylated derivatives of different pNP-sugars in a regioselective manner, as well as of some phenolic compounds of industrial interest, such as epigallocatechin gallate (EGCG). Conclusions In this work, we report the characterization of a novel glucotolerant 1,2-β-glucosidase, which also has a considerable activity on 1,4-β-glucosyl bonds, that has been cloned in P. pastoris, produced, purified and characterized. In addition, the enzyme was converted into an efficient glycosynthase, able to transfer glucose molecules to a diversity of acceptors for obtaining compounds of interest. The remarkable capacities of BGL-1 and its glycosynthase mutant, both in hydrolysis and synthesis, suggest that it could be an interesting tool for biotechnological applications.

show abstract

Section: Discussionmentioning

confidence: 79%

A glucotolerant β-glucosidase from the fungus Talaromyces amestolkiae and its conversion into a glycosynthase for glycosylation of phenolic compounds

et al. 2020

View full text Add to dashboard Cite

show abstract

“…Pichia pastoris has been used to heterologously express cellulolytic enzymes from the fungi Periconia sp. , Aspergillus fumigatus Z5 strain, Myceliophthora thermophila , Penicillium funiculosum NCL1 strain, Neurospora crassa and Talaromyce leycettanus , with K m values ranging from 0.18 to 2.5 mM for p NPG and 0.5 to 10.4 mM for cellobiose (Table 2) [14, 15, 17, 36–39]. Hence, D2-BGL expressed in P. pastoris has relatively high substrate affinity (i.e., low K m ).…”

Section: Resultsmentioning

confidence: 99%

Chaetomella raphigera β-glucosidase D2-BGL has intriguing structural features and a high substrate affinity that renders it an efficient cellulase supplement for lignocellulosic biomass hydrolysis

Kao

Kuo

Lee

et al. 2019

Biotechnol Biofuels

View full text Add to dashboard Cite

Background: To produce second-generation biofuels, enzymatic catalysis is required to convert cellulose from lignocellulosic biomass into fermentable sugars. β-Glucosidases finalize the process by hydrolyzing cellobiose into glucose, so the efficiency of cellulose hydrolysis largely depends on the quantity and quality of these enzymes used during saccharification. Accordingly, to reduce biofuel production costs, new microbial strains are needed that can produce highly efficient enzymes on a large scale. Results: We heterologously expressed the fungal β-glucosidase D2-BGL from a Taiwanese indigenous fungus Chaetomella raphigera in Pichia pastoris for constitutive production by fermentation. Recombinant D2-BGL presented significantly higher substrate affinity than the commercial β-glucosidase Novozyme 188 (N188; K m = 0.2 vs 2.14 mM for p-nitrophenyl β-d-glucopyranoside and 0.96 vs 2.38 mM for cellobiose). When combined with RUT-C30 cellulases, it hydrolyzed acid-pretreated lignocellulosic biomasses more efficiently than the commercial cellulase mixture CTec3. The extent of conversion from cellulose to glucose was 83% for sugarcane bagasse and 63% for rice straws. Compared to N188, use of D2-BGL halved the time necessary to produce maximal levels of ethanol by a semi-simultaneous saccharification and fermentation process. We upscaled production of recombinant D2-BGL to 33.6 U/mL within 15 days using a 1-ton bioreactor. Crystal structure analysis revealed that D2-BGL belongs to glycoside hydrolase (GH) family 3. Removing the N-glycosylation N68 or O-glycosylation T431 residues by site-directed mutagenesis negatively affected enzyme production in P. pastoris. The F256 substrate-binding residue in D2-BGL is located in a shorter loop surrounding the active site pocket relative to that of Aspergillus β-glucosidases, and this short loop is responsible for its high substrate affinity toward cellobiose. Conclusions: D2-BGL is an efficient supplement for lignocellulosic biomass saccharification, and we upscaled production of this enzyme using a 1-ton bioreactor. Enzyme production could be further improved using optimized fermentation, which could reduce biofuel production costs. Our structure analysis of D2-BGL offers new insights into GH3 β-glucosidases, which will be useful for strain improvements via a structure-based mutagenesis approach.

show abstract

“…The effects of temperature and pH on Lb0241 activity were determined according to the method of Li et al (2018) with slight modifications. The effect of pH on Lb0241 activity was determined in 500 mM citric acid–Na 2 HPO 4 phosphate buffer with various pH values ranging from 1.5 to 8.0 at 37°C for 30 min.…”

Section: Methodsmentioning

confidence: 99%

Cloning and expression of Lactobacillus brevis β‐glucosidase and its effect on the aroma of strawberry wine

Xia

Wang

et al. 2022

Food Processing Preservation

View full text Add to dashboard Cite

Aroma is one of the key characteristics of fruit wine and is correlated to volatile organic compounds, which greatly influences the purchase intention of consumers (Zhang, Wang, et al., 2021). Although the major aroma compounds, mainly derived from the fruit, are produced during the alcoholic fermentation stage, a number of odorless non-volatile glycosides are also present, which are often considered as aroma precursors; the hydrolysis of these glycosides could significantly improve the natural aroma compounds of fruit wine (Aaby et al., 2018;Michlmayr et al., 2010). Therefore, researchers have attempted to develop effective methods to hydrolyze these nonvolatile glycosides to release the free aroma compounds (Michlmayr et al., 2010). Lactic acid bacteria (LAB) can hydrolyze glycosylated

show abstract

A Novel Thermostable GH3β-Glucosidase fromTalaromyce leycettanuswith Broad Substrate Specificity and Significant Soybean Isoflavone Glycosides-Hydrolyzing Capability

Cited by 18 publications

References 31 publications

A glucotolerant β-glucosidase from the fungus Talaromyces amestolkiae and its conversion into a glycosynthase for glycosylation of phenolic compounds

A glucotolerant β-glucosidase from the fungus Talaromyces amestolkiae and its conversion into a glycosynthase for glycosylation of phenolic compounds

Chaetomella raphigera β-glucosidase D2-BGL has intriguing structural features and a high substrate affinity that renders it an efficient cellulase supplement for lignocellulosic biomass hydrolysis

Cloning and expression of Lactobacillus brevis β‐glucosidase and its effect on the aroma of strawberry wine

Contact Info

Product

Resources

About