Three amino acid changes contribute markedly to the thermostability of β-glucosidase BglC from Thermobifida fusca

Pei, Xiaokun; Yi, Zhuolin; Tang, Chuan-Gen; Wu, Zhong‐Liu

doi:10.1016/j.biortech.2010.11.025

Cited by 45 publications

(29 citation statements)

References 31 publications

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“…Such characteristic of mesophilic enzymes with excellent thermostability is usually desired because it could avoid the necessity of applying high reaction temperature when using thermophilic enzymes to reduce energy consumption, as well as to fit the thermal tolerance of other reactants, while at the same time, maintain many advantages of a stable enzyme, such as less enzyme loading, easy catalyst manipulation, and flexible process operations [42][43][44].…”

Section: Discussionmentioning

confidence: 99%

Identification and characterization of a novel “thermophilic-like” Old Yellow Enzyme from the genome of Chryseobacterium sp. CA49

Pei

2014

Journal of Molecular Catalysis B: Enzymatic

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Section: Discussionmentioning

confidence: 99%

Identification and characterization of a novel “thermophilic-like” Old Yellow Enzyme from the genome of Chryseobacterium sp. CA49

Pei

2014

Journal of Molecular Catalysis B: Enzymatic

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“…2B). In general, an increase in the number of hydrogen bonds between side chains of residues improves protein thermostability (22,25). This suggests that the higher number in putative hydrogen bonds in the S213C variant enzyme may contribute to its increased thermostability.…”

Section: Screening Of Thermostable Variants Created By Random Mutagenmentioning

confidence: 99%

Improvement in the Thermostability of d -Psicose 3-Epimerase from Agrobacterium tumefaciens by Random and Site-Directed Mutagenesis

Choi

Yeom

et al. 2011

Appl Environ Microbiol

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The S213C, I33L, and I33L S213C variants of D-psicose 3-epimerase from Agrobacterium tumefaciens, which were obtained by random and site-directed mutagenesis, displayed increases of 2.5, 5, and 7.5°C in the temperature for maximal enzyme activity, increases of 3.3-, 7.2-, and 29.9-fold in the half-life at 50°C, and increases of 3.1, 4.3, and 7.6°C in apparent melting temperature, respectively, compared with the wild-type enzyme. Molecular modeling suggests that the improvement in thermostability in these variants may have resulted from increased putative hydrogen bonds and formation of new aromatic stacking interactions. The immobilized wild-type enzyme with and without borate maintained activity for 8 days at a conversion yield of 70% (350 g/liter psicose) and for 16 days at a conversion yield of 30% (150 g/liter psicose), respectively. After 8 or 16 days, the enzyme activity gradually decreased, and the conversion yields with and without borate were reduced to 22 and 9.6%, respectively, at 30 days. In contrast, the activities of the immobilized I33L S213C variant with and without borate did not decrease during the operation time of 30 days. These results suggest that the I33L S213C variant may be useful as an industrial producer of D-psicose.D-Psicose (D-allulose), a carbon-3 epimer of D-fructose, is present in small quantities as a nonfermentable constituent of cane molasses (1), as a sugar moiety of the nucleoside antibiotic psicofuranine (4), and as a free sugar in wheat (21) and Itea plants (5). This rare sugar provides suitable sweetness, smooth texture, favorable mouthfeel, and long-time storage stability in food products (23). Psicose is considered to be a potential reduced energy sweetener because the sugar suppresses hepatic lipogenic enzyme activity (18) and does not contribute to calorie production (19).Biological production of psicose from fructose has been studied using D-psicose 3-epimerase from Agrobacterium tumefaciens (10, 13, 16) and D-tagatose 3-epimerases from Pseudomonas cichorii (7,24) and Rhodobacter sphaeroides (26). D-Psicose 3-epimerase from A. tumefaciens has been more effective than the D-tagatose 3-epimerases for the production of psicose. However, D-psicose 3-epimerase is inefficient for the industrial production of psicose because of its short half-life of 63 min at 50°C (10). Thus, improvement in the thermostability of A. tumefaciens D-psicose 3-epimerase is essential for the industrial production of psicose.Random mutagenesis and rational protein design are typical tools for improving enzyme thermostability in the protein engineering field. Random mutagenesis techniques such as errorprone PCR and DNA shuffling can be easily applied for the improvement of enzyme thermostability (8, 9, 12, 28) because these techniques do not require detailed structural information or accurate predictions at the substituted residues (14).In the present study, thermostable variants of D-psicose 3-epimerase from A. tumefaciens were obtained by random and site-directed mutagenesis. The temperature fo...

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“…This enzyme is commonly produced by a wide range of organisms, including bacteria, fungi, and plants. β-Glucosidases play pivotal roles in various biotechnological processes such as the reduction of cellobiose accumulation by the action of exo-glucanases to minimize the end-product inhibition [2] and the modification of cyanogenesis, glycolipids, and secondary metabolites [1]. They are involved in the bioconversion of lignocellulose to glucose, producing hydrogen with other exo-or endo-hydrolases and using glucose as an ingredient for fermentation [3,4].…”

Section: Introductionmentioning

confidence: 99%

Over-Expression of the Thermobifida fusca β-Glucosidase in a Yarrowia lipolytica Transformant to Degrade Soybean Isoflavones

et al. 2018

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Abstract:A gene (bgl) encoding a β-glucosidase in thermophilic actinomycete Thermobifida fusca NTU 22 was cloned into a Yarrowia lipolytica expression system. Heterologous expression resulted in extracellular β-glucosidase production with activity as high as 630 U/mL in a Hinton flask culture filtrate. This recombinant β-glucosidase was purified 9.2-fold from crude culture filtrate by DEAE-Sepharose FF column chromatography as measured by its increase in specific activity. The overall yield of the purified enzyme was 47.5%. The molecular weight of the purified β-glucosidase estimated by SDS-PAGE was 45 kDa, which agreed with the predicted molecular weight based on the nucleotide sequence. About 15% enzyme activity loss was observed after the enzyme was heat-treated at 50 • C for 180 min. It was also found that the activity of the enzyme was inhibited by Hg 2+ , Cu 2+ , Ba 2+ , Ag + , p-chloromercuribenzene, and iodoacetate. The β-glucosidase from T. fusca had the most activity for daidzein-7-glucoside and genistein-7-glucoside among the tested flavonoid glycosides, but there was moderate or little activity for luteolin-7-glucoside, cyanidine-3-glucoside, and quercetin-3-glucoside. These properties are important for the soybean isoflavone applications of this β-glucosidase.

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Three amino acid changes contribute markedly to the thermostability of β-glucosidase BglC from Thermobifida fusca

Cited by 45 publications

References 31 publications

Identification and characterization of a novel “thermophilic-like” Old Yellow Enzyme from the genome of Chryseobacterium sp. CA49

Identification and characterization of a novel “thermophilic-like” Old Yellow Enzyme from the genome of Chryseobacterium sp. CA49

Improvement in the Thermostability of d -Psicose 3-Epimerase from Agrobacterium tumefaciens by Random and Site-Directed Mutagenesis

Over-Expression of the Thermobifida fusca β-Glucosidase in a Yarrowia lipolytica Transformant to Degrade Soybean Isoflavones

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