Dataset Information

Characterization of a Novel Hyperthermophilic GH1 β-Glucosidase from Acidilobus sp. and Its Application in the Hydrolysis of Soybean Isoflavone Glycosides.

ABSTRACT: A putative β-glucosidase gene, BglAc, was amplified from Acidilobus sp. through metagenome database sampling from a hot spring in Yellowstone National Park. BglAc is composed of 485 amino acid residues and bioinformatics analysis showed that it belongs to the GH1 family of β-glucosidases. The gene was successfully expressed in Escherichia coli with a molecular weight of approximately 55.3 kDa. The purified recombinant enzyme showed the maximum activity using p-nitrophenyl-β-D-glucopyranoside (pNPG) as the substrate at optimal pH 5.0 and 100 °C. BglAc exhibited extraordinary thermostability, and its half-life at 90 °C was 6 h. The specific activity, K_m, V_max, and K_cat/K_m of BglAc toward pNPG were 357.62 U mg^-1, 3.41 mM, 474.0 μmol min^-1·mg^-1, and 122.7 s^-1mM^-1. BglAc exhibited the characteristic of glucose tolerance, and the inhibition constant Ki was 180.0 mM. Furthermore, a significant ethanol tolerance was observed, retaining 96% relative activity at 10% ethanol, and even 78% at 20% ethanol, suggesting BglAc as a promising enzyme for cellulose saccharification. BglAc also had a strong ability to convert the major soybean isoflavone glycosides (daidzin, genistin, and glycitin) into their corresponding aglycones. Overall, BglAc was actually a new β-glucosidase with excellent thermostability, ethanol tolerance, and glycoside hydrolysis ability, indicating its wide prospects for applications in the food industry, animal feed, and lignocellulosic biomass degradation.

SUBMITTER: He J

PROVIDER: S-EPMC10974336 | biostudies-literature | 2024 Mar

REPOSITORIES: biostudies-literature

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Characterization of a Novel Hyperthermophilic GH1 β-Glucosidase from Acidilobus sp. and Its Application in the Hydrolysis of Soybean Isoflavone Glycosides.

He Jinjian J Li Yuying Y Sun Xihang X Zuo Dinghui D Wang Mansheng M Zheng Xia X Yu Pinglian P Shi Pengjun P

Microorganisms 20240307 3

A putative β-glucosidase gene, BglAc, was amplified from Acidilobus sp. through metagenome database sampling from a hot spring in Yellowstone National Park. BglAc is composed of 485 amino acid residues and bioinformatics analysis showed that it belongs to the GH1 family of β-glucosidases. The gene was successfully expressed in Escherichia coli with a molecular weight of approximately 55.3 kDa. The purified recombinant enzyme showed the maximum activity using p-nitroph ...[more]

PMID: 38543584

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Project description:A novel hyperthermophilic, anaerobic filamentous archaeon, Thermofilum adornatum strain 1910bT, is capable of growing with cellulose as its sole carbon and energy source. This strain was isolated from a terrestrial hot spring in Kamchatka, Russia. The isolate 1910bT grew optimally at a temperature of 80°C and a pH of 5.5-6.0, producing cell-bound inducible cellulases. During genome analysis, genes, encoding various glycosidases (GHs) involved in oligo- and polysaccharide hydrolysis and genes for the fermentation of sugars were identified. No homologs of currently known cellulase families were found among the GHs encoded by the 1910bT genome, suggesting that novel proteins are involved. To figure this out, a proteomic analysis of cells grown on cellulose or pyruvate (as a control) was performed. Both in-depth genomic and proteomic analyses revealed four proteins (Cel25, Cel30, Cel40, and Cel45) that were the most likely to be involved in the cellulose hydrolysis in this archaeon. Two of these proteins (Cel30 and Cel45) were hypothetical according to genome analysis, while the other two (Cel25 and Cel40) have GH3 and GH1 domains, respectively. The respective genes were heterologously expressed in Escherichia coli BL21 (DE3), and enzymatic activities of recombinant proteins were measured with carboxymethyl cellulose (CMC), Avicel and cellobiose as substrates. It was revealed that the Cel30 and Cel25 proteins were likely exoglucanases with side beta-glucosidase and endoglucanase activities, that Cel40 was a multifunctional glucanase capable of hydrolyzing beta-1,4-glucosides of various lengths, and that Cel45 was an endoglucanase with side exoglucanase activity. Taking into account that the cellulolytic activity of T. adornatum 1910bT surface protein fractions was inducible, that recombinant Cel25 and Cel30 were much less active than Cel40 and Cel45, and that their gene expressions were (almost) non-induced by CMC, we suggest that Cel40 and Cel45 play a major role in the degradation of cellulose, while Cel25 and Cel30 act only as accessory enzymes.

Dataset Information

Characterization of a Novel Hyperthermophilic GH1 β-Glucosidase from Acidilobus sp. and Its Application in the Hydrolysis of Soybean Isoflavone Glycosides.

Publications

Characterization of a Novel Hyperthermophilic GH1 β-Glucosidase from <i>Acidilobus</i> sp. and Its Application in the Hydrolysis of Soybean Isoflavone Glycosides.

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