Purification of an l-arabinose isomerase from Enterococcus faecium DBFIQ E36 employing a biospecific affinity strategy

Torres, Pedro; Manzo, Ricardo Martín; Rubiolo, Amelia Catalina; Batista‐Viera, Francisco; Mammarella, E

doi:10.1016/j.molcatb.2014.01.023

Cited by 16 publications

(15 citation statements)

References 49 publications

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“…The production of d -tagatose from d -galactose through enzymatic synthesis is attractive as an industrial process [ 7 , 8 ] when compared with the chemical conversion process, i.e., mild reaction conditions, no by-products, a lack of salt in the produced wastewater, and the possibility of recycling the unreacted d -galactose [ 9 ]. l -Arabinose isomerase (EC 5.3.1.4) ( l -AI) encoded by araA -like genes has been the most studied enzyme for the conversion of d -galactose to d -tagatose due to the industrial viability of this process, since d -galactose can be obtained from the lactose contained in cheese whey as a cheap substrate [ 10 , 11 ]. For this reason, the production of d -tagatose from d -galactose using l -AI has been studied in recent years [ 6 , 11 , 12 , 13 ].…”

Section: Introductionmentioning

confidence: 99%

Engineering the l-Arabinose Isomerase from Enterococcus Faecium for d-Tagatose Synthesis

Sousa

Manzo

Garcı́a³

et al. 2017

Molecules

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l-Arabinose isomerase (EC 5.3.1.4) (l-AI) from Enterococcus faecium DBFIQ E36 was overproduced in Escherichia coli by designing a codon-optimized synthetic araA gene. Using this optimized gene, two N- and C-terminal His-tagged-l-AI proteins were produced. The cloning of the two chimeric genes into regulated expression vectors resulted in the production of high amounts of recombinant N-His-l-AI and C-His-l-AI in soluble and active forms. Both His-tagged enzymes were purified in a single step through metal-affinity chromatography and showed different kinetic and structural characteristics. Analytical ultracentrifugation revealed that C-His-l-AI was preferentially hexameric in solution, whereas N-His-l-AI was mainly monomeric. The specific activity of the N-His-l-AI at acidic pH was higher than that of C-His-l-AI and showed a maximum bioconversion yield of 26% at 50 °C for d-tagatose biosynthesis, with Km and Vmax parameters of 252 mM and 0.092 U mg−1, respectively. However, C-His-l-AI was more active and stable at alkaline pH than N-His-l-AI. N-His-l-AI follows a Michaelis-Menten kinetic, whereas C-His-l-AI fitted to a sigmoidal saturation curve.

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Section: Introductionmentioning

confidence: 99%

Engineering the l-Arabinose Isomerase from Enterococcus Faecium for d-Tagatose Synthesis

Sousa

Manzo

Garcı́a³

et al. 2017

Molecules

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“…Due to numerous health benefits of d ‐tagatose, an extensive research is going on across the world to develop the efficient process for its production after due contemplation to food and environmental safety, human health, and consumer awareness. The biological conversion of d ‐galactose into d ‐tagatose by application of l‐ AI enzyme seems to be a viable process because it relies on isomerisation of d ‐galactose, an industrial by‐product of cheese industry (Torres, Manzo, Rubiolo, Batista‐Vieraa, & Mammarella, ). During the past few decades, biological production of d ‐tagatose has been studied using l‐ AI from E. coli (Yoon, Kim, & Oh, ), Lcatobacillus fermentum (Xu et al., ), Bacillus coagulans (Mei, Wang, Zang, Zheng, & Ouyang, ), Geobacillus thermodenitrificans (Kim & Oh, ), Geobacillus sterothermophilus (Lee et al., ), Thermotoga neapolitana (Kim et al., ), Thermotoga maritime (Lee et al., ), and Anoxybacillus flavithermus (Li, Zhu, Liu, & Sun, ).…”

Section: L‐arabinose Isomerasementioning

confidence: 99%

Tagatose as a Potential Nutraceutical: Production, Properties, Biological Roles, and Applications

Roy

Chikkerur

Roy

et al. 2018

Journal of Food Science

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Nutraceuticals are gaining importance owing to their potential applications in numerous sectors including food and feed industries. Among the emerging nutraceuticals, d‐tagatose occupies a significant niche because of its low calorific value, antidiabetic property and growth promoting effects on beneficial gut bacteria. As d‐tagatose is present in minute quantities in naturally occurring food substances, it is produced mainly by chemical or biological means. Recently, attempts were made for bio‐production of d‐tagatose using l‐arabinose isomerase enzyme to overcome the challenges of chemical process of production. Applications of d‐tagatose for maintaining health and wellbeing are increasing due to growing consumer awareness and apprehension against modern therapeutic agents. This review outlines the current status on d‐tagatose, particularly its production, properties, biological role, applications, and the future perspectives.

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“…Pure preparations of the enzyme were achieved through clarification by membrane filtration, ammonium sulfate precipitation (85 % saturation), followed by l-arabitol-Sepharose CL-4B affinity chromatography, according to Torres et al [56].…”

Section: Enzyme Production and Purificationmentioning

confidence: 99%

“…Analysis of oxirane groups was performed according to Sundberg and Porath [54] with several modifications proposed by Torres et al [56].…”

Section: Determination Of Oxirane Group Content On Activated Supportsmentioning

confidence: 99%

“…For the current research, L-AI from Enterococcus faecium DBFIQ E36 strain, isolated from raw cow milk, was purified up to electrophoretical homogeneity and employed to obtain insoluble biocatalysts. Briefly, this enzyme presents a homo-tetrameric structure with a MW of 187 kDa determined by gel filtration, a pI of 3.8, and was also characterized as a Mn 2+ ion-activated enzyme [37,56]. Furthermore, the use of low-cost supports such as calcium carbonate, chitin and chitosan represents an economical advantage over commercial resins [11].…”

Section: Introductionmentioning

confidence: 99%

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Chemical improvement of chitosan-modified beads for the immobilization of Enterococcus faecium DBFIQ E36 l-arabinose isomerase through multipoint covalent attachment approach

Manzo

Sousa

Fenoglio

et al. 2015

Journal of Industrial Microbiology and Biotechnology

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D-tagatose is produced from D-galactose by the enzyme L-arabinose isomerase (L-AI) in a commercially viable bioprocess. An active and stable biocatalyst was obtained by modifying chitosan gel structure through reaction with TNBS, D-fructose or DMF, among others. This led to a significant improvement in L-AI immobilization via multipoint covalent attachment approach. Synthetized derivatives were compared with commercial supports such as Eupergit(®) C250L and glyoxal-agarose. The best chitosan derivative for L-AI immobilization was achieved by reacting 4 % (w/v) D-fructose with 3 % (w/v) chitosan at 50 °C for 4 h. When compared to the free enzyme, the glutaraldehyde-activated chitosan biocatalyst showed an apparent activity of 88.4 U g (gel) (-1) with a 211-fold stabilization factor while the glyoxal-agarose biocatalyst gave an apparent activity of 161.8 U g (gel) (-1) with an 85-fold stabilization factor. Hence, chitosan derivatives were comparable to commercial resins, thus becoming a viable low-cost strategy to obtain high active L-AI insolubilized derivatives.

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Purification of an l-arabinose isomerase from Enterococcus faecium DBFIQ E36 employing a biospecific affinity strategy

Cited by 16 publications

References 49 publications

Engineering the l-Arabinose Isomerase from Enterococcus Faecium for d-Tagatose Synthesis

Engineering the l-Arabinose Isomerase from Enterococcus Faecium for d-Tagatose Synthesis

Tagatose as a Potential Nutraceutical: Production, Properties, Biological Roles, and Applications

Chemical improvement of chitosan-modified beads for the immobilization of Enterococcus faecium DBFIQ E36 l-arabinose isomerase through multipoint covalent attachment approach

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