Scientific Reports
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2015
·Vol. 5
·期 1
·1-12
·
IF 5.578
Yang Yang,Zhang Xinxin,Yin Qiang,Fang Wei,Fang Zemin,Wang Xiaotang,Zhang Xuecheng,Xiao Yazhong
β-Glucosidases are enzymes that hydrolyze β-glycosidic bonds to release non-reducing terminal glucosyl residues from glycosides and oligosaccharides and thus have significant application potential in industries. However, most β-glucosidases are feedback inhibited by the glucose product, which restricts their application. Remarkably, some β-glucosidases of the glycoside hydrolase (GH) 1 family are tolerant to or even stimulated by glucose. Elucidation of the mechanisms of glucose tolerance and stimulation of the GH1 β-glucosidases will be crucial to improve their application through enzyme engineering. In this study, by comparing the primary and tertiary structures of two GH1 β-glucosidases with distinct glucose dependence, some putative glucose-dependence relevant sites were mutated to investigate their exact roles. Both biochemical and structural characterization of the mutants suggested that some sites at the entrance and middle of the substrate channel regulate the effects of glucose and the relative binding affinity/preference of these sites to glucose modulates the glucose dependence. A mechanism was therefore proposed to interpret the glucose dependence of GH1 β-glucosidases. This research provides fresh insight into our current understanding of the properties and mechanisms of GH1 β-glycosidases and related enzymes that modulate their activity via feedback control mechanism.
研究领域:
生物化学
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酶学
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生物技术
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DOI
PubMed
高影响因子
MOLECULES
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2024
·Vol. 29
·期 22
·5280
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IF 4.2
Xiaoling Zhang,Yitong Wang,Tiantian Zhang,Ziqiao Yuan,Yongjun Wei
Epimedium Folium(EF) is a traditional Chinese herbal medicine, and its primary bioactive ingredients, such as icariin, are flavonoid glycosides. A rare EF flavonoid, baohuoside I, exhibits superior bioactivities and enhanced bioavailability compared to its metabolic precursor icariin. The biotransformation of icariin to baohuoside I can be effectively and specifically achieved by β-glucosidases. In this study, 33 candidate full-length β-glucosidase genes were screened from a previously built carbohydrate active enzyme (CAZyme) gene dataset derived from cow fecal microbiota. Thirteen of them exhibited β-glucosidase activity, with DCF-bgl-26 and DCF-bgl-27 showing relatively high expression levels and β-glucosidase activity. The maximum β-glucosidase activity of DCF-bgl-26 and DCF-bgl-27 was achieved at 45 °C and pH 6.0, with DCF-bgl-26 demonstrating better thermostability and pH tolerance compared to DCF-bgl-27. The activities of DCF-bgl-26 and DCF-bgl-27 were 123.2 U/mg protein and 157.9 U/mg protein, respectively, both of which are higher than those of many bacterial β-glucosidases. Structure analysis suggested that both β-glucosidases possess canonical (β/α)8-TIM barrel fold structure of GH1 family β-glucosidases. Thin-layer chromatography results showed that both enzymes could efficiently convert icariin to baohuoside I in 30 min, indicating they have potential application in the production of high value rare baohuoside I.
研究领域:
生物化学
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生物技术
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药理学
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微生物学
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DOI
PubMed
高影响因子
Molecular Catalysis
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2024
·
IF 3.9
Qiqi Xing,Xiaoyu Wang,Qi Li,Changning Lu,Yi Wu,Jianjun Pei,Linguo Zhao
β-Galactosidase is a crucial class of biocatalysts, renowned for its ability to catalyze the hydrolysis of non-reducing terminal β-d-galactoside bonds, while also demonstrating significant potential in galactosylation synthesis reactions. In this study, a novel GH1 β-galactosidase gene VthBga was screened and cloned from Vulcanisaeta thermophila . Soluble expression of the VthBga was induced at 47 °C in Escherichia coli BL21. VthBga exhibited optimal activity at 95 °C and pH 4.5 with p -nitrophenyl-β-d-galactoside ( p NPGal) as a substrate and displayed excellent thermotolerance below 90 °C. In addition, VthBga possessed not only a diverse array of substrate hydrolytic activities but also inherent galactosylation activity, rendering it a promising candidate for the galactosylation of the natural compound. To enhance its transglycosylation activity, a superior mutant Vth-A266G/F441T was obtained by adjusting the transglycosylation/hydrolysis ratio through a semi-rational design. Under optimal conditions, this mutant produced 44.5 % tyrosol galactoside, representing a significant 2.3-fold increase compared to the wild-type strain, highlighting its practical value in related biosynthesis applications.
研究领域:
生物化学
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分子生物学
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酶工程
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高影响因子
ANTONIE VAN LEEUWENHOEK INTERNATIONAL JOURNAL OF GENERAL AND MOLECULAR MICROBIOLOGY
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2020
·Vol. 113
·期 10
·1467-1477
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IF 2.674
Yin Yi-Rui,Sang Peng,Yang Feng-Li,Li Tao,Yang Run-Feng,Liu Hong-Yan,Luo Zong-Long,Li Wen-Jun,Yang Li-Quan
A β-glucosidase gene ( bsbgl1a ) from Bacillus sp. CGMCC 1.16541 was expressed in Escherichia coli BL21 and subsequently characterized. The amino acid sequence shared 83.64% identity with β-glucosidase (WP_066390903.1) from Fictibacillus phosphorivorans . The recombinant β-glucosidase (BsBgl1A) had a molecular weight of 52.2 kDa and could hydrolyze cellobiose, cellotriose, cellotetrose, p-nitrophenyl-β- d -glucopyranoside ( p NPG), and p-nitrophenyl-β- d -xylopyranoside ( p NPX). Optimal activity for BsBgl1A was recorded at 45 °C with a pH between 5.6 and 7.6, and 100% of its activity was maintained after a 24 h incubation between pH 4 and 9. Kinetic characterization revealed an enzymatic turnover (Kcat) of 616 ± 2 s −1 (with cellobiose) and 3.5 ± 0.1 s −1 (with p-nitrophenyl-β- d -glucopyranoside). Interestingly, the recombinant enzyme showed cupric ion (Cu 2+ ), sodium dodecyl sulfate (SDS) and alcohol tolerance at 10 mM for Cu 2+ and 10% for both SDS and alcohol. Additionally, BsBgl1A had high tolerance for glucose (Ki = 2095 mM), which is an extremely desirable feature for industrial applications. Following the addition of BsBgl1A (0.05 mg/ml) to a commercial cellulase reaction system, glucose yields from sugarcane bagasse increased 100% after 1 day at 45 °C. This work identifies a Cu 2+ , SDS, alcohol, and glucose tolerant GH1 β-glucosidase with potential applications in the hydrolysis of cellulose for the bioenergy industry.
研究领域:
酶学
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微生物学
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生物技术
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生物化学
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DOI
PubMed
高影响因子
Biomass Conversion and Biorefinery
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2020
·Vol. 11
·期 4
·1245-1253
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IF 2.602
Yin Yi-Rui,Sang Peng,Xiao Min,Xian Wen-Dong,Dong Zhou-Yan,Liu Lan,Yang Li-Quan,Li Wen-Jun
A β-glucosidase obtained from Thermobifida halotolerans YIM 90462 T was expressed in Escherichia coli BL21 and subsequently characterized. The recombinant enzyme ThBGL1A showed optimal activity at 45 °C and pH 6, but also showed high activity from 40 to 55 °C and pH 5.6–6.6. Its half-life activity was 58 min at 50 °C. ThBGL1A exhibited notable cold-adapted activity, retaining 13.1%, 49.4%, and 83.5% of its optimal activity at 5, 25, and 30 °C, respectively. Kinetic characterization revealed an enzymatic turnover (Kcat) of 30 s −1 (cellobiose), 41.8 s −1 (p-nitrophenyl-β- d -glucopyranoside), and 52.6 s −1 (p-nitrophenyl-β- d -galactopyranoside). Moreover, ThBGL1A had high tolerance for salt, xylose, and glucose, which are extremely desirable features for industrial applications. Interestingly, its Ki for glucose was 932 mM and more than 80% of its optimal activity in the presence of 2000 mM xylose. After the addition of ThBGL1A (0.05 mg/ml) to a commercial cellulase reaction system, glucose yields from sugarcane bagasse were increased 20% and 18% after 1 day at 30 °C and 45 °C, respectively. Overall, this work identifies a cold-adapted, salt- and glucose-tolerant β-glucosidase with potential applications in commercial cellulose utilization and the bioenergy industry.
研究领域:
生物化学
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生物技术
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酶学
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可持续能源
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DOI
高影响因子
APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
·
2020
·Vol. 104
·期 11
·4927-4943
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IF 3.53
Deng Pengjun,Meng Chunyu,Wu Yan,Xu Jie,Tang Xiaoqing,Zhang Xuecheng,Xiao Yazhong,Wang Xiaotang,Fang Zemin,Fang Wei
A novel β -glucosidase, BglD1 with high β -galactosidase and transglycosidation activities, was screened and cloned from the deep-sea bacterium Bacillus sp. D1. BglD1 exhibited the maximal β -glucosidase and β -galactosidase activities at 55–60 °C and pH 5.5–6.0. The enzyme maintained approximately 50% of its original activity at 35 °C and pH 6.0 after 120-h incubation. When applied to synthesize galacto-oligosaccharides (GOS), BglD1 generated 118.3 g/L GOS (33.8% (w/w)) from 350 g/L lactose, with trisaccharide Gal- β (1 → 3)-Lac and disaccharide Gal- β (1 → 4)-Gal as the main components. Furthermore, BglD1 could hydrolyze lactose in milk and produce GOS simultaneously. Using milk as the substrate, BglD1 hydrolyzed 88.5% lactose and produced 3.3 g/L GOS after incubation at 30 °C for 1 h. To improve the transglycosidation activity, a mutant BglD1:E224T was generated based on the semi-rational design. The GOS yield of BglD1:E224T was 11.5% higher than that of BglD1 when using lactose solution as the substrate. Thus, BglD1 and the mutant could be used as beneficial alternatives of the existing β -galactosidases for the production of GOS.
研究领域:
微生物学
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酶工程
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食品科学
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DOI
PubMed
高影响因子
Microorganisms
·
2025
·Vol. 13
·期 9
·2033
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IF 4.2
Yanmei Cai,Xinyu Li,Shuang Chen,Qichao Liu,Hongxiang Lu,Jiahui Xie,Wei Li,Guiying Chen
Microorganisms with chitin-degrading capabilities play a crucial role in the biological control of crop pests and diseases as well as in the treatment of organic waste. In this study, a chitin-degrading bacterium, designated L2-2, was isolated from the intestine ofOdorrana margaretaecollected in Mount Emei, Sichuan, China. Based on physiological and biochemical characteristics, 16S rRNA gene sequencing, and phylogenetic analysis of 31 conserved housekeeping genes in the whole genome, strain L2-2 was identified as a member of the genusRoseateles, namedRoseatelessp. L2-2. This strain is able to grow on agar medium with colloidal chitin as the sole carbon source and form clear hydrolysis zones. After optimizing fermentation conditions (including concentrations of nitrogen and carbon sources, culture time, and pH), the enzyme activity was increased to 3.46 U/mL, which was 24 times higher than the initial enzyme activity. Functional genome annotation showed that the strain contains genes encoding endochitinases of the GH18, GH23, and GH46 families, as well as genes encoding β-glucosidases of the GH1, GH2, GH3, and GH109 families, indicating its genetic basis for chitin-degrading potential. This study expands the diversity of known chitin-degrading bacteria and provides a promising microbial resource for the bioremediation of chitinous waste and sustainable pest control in agriculture.
研究领域:
微生物学
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环境生物技术
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分子生物学
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应用微生物学
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生物修复
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农业微生物学
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PubMed
高影响因子
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