Molecular Cloning of the Amylosucrase Gene from a Moderate Thermophilic Bacterium Deinococcus Geothermalis and Analysis of its Dual Enzyme Activity

Seo, Dong-Ho; Jung, Jong‐Hyun; Ha, Suk‐Jin; Yoo, Sang‐Ho; Kim, Tae-Jip; Cha, Jaeho; Park, Cheon-Seok

doi:10.1533/9781845695750.1.125

Cited by 24 publications

(39 citation statements)

References 32 publications

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“…Construction of the inducible DGAS expression vector, pGEX-DGAS, was described previously (Seo et al 2008). The dgas gene was obtained from the plasmid DNA of pGEX-DGAS.…”

Section: Construction Of Constitutive Expression Vectormentioning

confidence: 99%

“…Two oligonucleotide primers containing NdeI and XhoI recognition sites (underline), pHCDGAS-F (5′-CATATGCTGAAA G A C G T G C T C A C T-3 ′ ) and pHCDGAS-R (5′ -CTCGAGTGCTGGAGCCTCCCCGGCGGT-3′), were used to amplify the dgas gene from pGEX-DGAS. PCR conditions were described previously (Seo et al 2008). The resulting PCR products were directly cloned into the pGEM-T Easy vector (Promega) followed by DNA sequencing analysis to confirm the absence of PCR-introduced errors.…”

Section: Construction Of Constitutive Expression Vectormentioning

confidence: 99%

“…This enzyme cleaves the glycosidic linkages of sucrose and transfers a glucose molecule from sucrose to the 4-positions of the non-reducing end of an α-1→4 glucan chain of the acceptor molecule, thereby creating an α-1→4 linked glucan polymer (Buttcher et al 1997;Potocki de Montalk et al 2000;Skov et al 2001;PotockiVeronese et al 2005). Recently, it was found that ASase could utilize molecules other than α-1→4 glucan as an acceptor; these molecules include galactose, xylose, salicin, and β-Ab (Seo et al 2008;Jung et al 2009;Seo et al 2009a). ASase genes from Neisseria polysaccharea, Deinococcus radiodurans, Deinococcus geothermalis (DGAS), and Alteromonas macleodii have been cloned and expressed in Escherichia coli (De Montalk et al 1999;Pizzut-Serin et al 2005;Seo et al 2008;Ha et al 2009).…”

Section: Introductionmentioning

confidence: 99%

“…Recently, it was found that ASase could utilize molecules other than α-1→4 glucan as an acceptor; these molecules include galactose, xylose, salicin, and β-Ab (Seo et al 2008;Jung et al 2009;Seo et al 2009a). ASase genes from Neisseria polysaccharea, Deinococcus radiodurans, Deinococcus geothermalis (DGAS), and Alteromonas macleodii have been cloned and expressed in Escherichia coli (De Montalk et al 1999;Pizzut-Serin et al 2005;Seo et al 2008;Ha et al 2009). These functional enzymes were employed to make a variety of attractive biomaterials including amylose-like polymer, resistant starch, dendritic nanoparticles, and biologically active compounds Putaux et al 2006;Jung et al 2009;Seo et al 2009a;Ryu et al 2010;Cho et al 2011).…”

Section: Introductionmentioning

confidence: 99%

“…In previous studies, all recombinant ASases were expressed using an inducible tac promoter in pGEX-based vectors (Pizzut-Serin et al 2005;Seo et al 2008;Ha et al 2009). In this system, the recombinant ASases were expressed as glutathione S-transferase (GST)-tagged fusion proteins and, therefore, the expressed enzymes had to be cut by special proteases to eliminate the large GST.…”

Section: Introductionmentioning

confidence: 99%

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High-yield enzymatic bioconversion of hydroquinone to α-arbutin, a powerful skin lightening agent, by amylosucrase

Seo

Jung

et al. 2012

Appl Microbiol Biotechnol

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α-Arbutin (α-Ab) is a powerful skin whitening agent that blocks epidermal melanin biosynthesis by inhibiting the enzymatic oxidation of tyrosine and L-3,4-dihydroxyphenylalanine (L-DOPA). α-Ab was effectively synthesized from hydroquinone (HQ) by enzymatic biotransformation using amylosucrase (ASase). The ASase gene from Deinococcus geothermalis (DGAS) was expressed and efficiently purified from Escherichia coli using a constitutive expression system. The expressed DGAS was functional and performed a glycosyltransferase reaction using sucrose as a donor and HQ as an acceptor. The presence of a single HQ bioconversion product was confirmed by thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC). The HQ bioconversion product was isolated by silica gel open column chromatography and its chemical structure determined by 1H and 13C nuclear magnetic resonance (NMR). The product was determined to be hydroquinone-O-α-D-glucopyranoside with a glucose molecule linked to HQ through an α-glycosidic bond. However, the production yield of the transfer reaction was significantly low (1.3%) due to the instability of HQ in the reaction mixture. The instability of HQ was considerably improved by antioxidant agents, particularly ascorbic acid, implying that HQ is labile to oxidation. A maximum yield of HQ transfer product of 90% was obtained at a 10:1 molar ratio of donor (sucrose) and acceptor (HQ) molecules in the presence of 0.2 mM ascorbic acid.

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“…Construction of the inducible DGAS expression vector, pGEX-DGAS, was described previously (Seo et al 2008). The dgas gene was obtained from the plasmid DNA of pGEX-DGAS.…”

Section: Construction Of Constitutive Expression Vectormentioning

confidence: 99%

Section: Construction Of Constitutive Expression Vectormentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

High-yield enzymatic bioconversion of hydroquinone to α-arbutin, a powerful skin lightening agent, by amylosucrase

Seo

Jung

et al. 2012

Appl Microbiol Biotechnol

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Amylosucrase: A Versatile Sucrose-Utilizing Transglucosylase for Glycodiversification

Tian

Chen

Zhang

2021

Novel Enzymes for Functional Carbohydrates Production

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Enzymatic modification of daidzin using heterologously expressed amylosucrase in Bacillus subtilis

Kim

Rha

Jung

et al. 2018

Food Sci Biotechnol

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Amylosucrases (ASase, EC 2.4.1.4) from Deinococcus geothermalis (DGAS) and Neisseria polysaccharea (NPAS) were heterologously expressed in Bacillus subtilis. While DGAS was successfully expressed, NPAS was not. Instead, NPAS was expressed in Escherichia coli. Recombinant DGAS and NPAS were purified using nickel-charged affinity chromatography and employed to modify daidzin to enhance its water solubility and bioavailability. Analyses by LC/MS revealed that the major products of transglycosylation using DGAS were daidzein diglucoside and daidzein triglucoside, whereas that obtained by NPAS was only daidzein diglucoside. The optimal bioconversion conditions for daidzein triglucoside, which was predicted to have the highest water-solubility among the daidzin derivatives, was determined to be 4% (w/v) sucrose and 250 mg/L daidzin in sodium phosphate pH 7.0, with a reaction time of 12 h. Taken together, we suggest that the yield and product specificity of isoflavone daidzin transglycosylation may be modulated by the source of ASase and reaction conditions.

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Molecular Cloning of the Amylosucrase Gene from a Moderate Thermophilic Bacterium Deinococcus Geothermalis and Analysis of its Dual Enzyme Activity

Cited by 24 publications

References 32 publications

High-yield enzymatic bioconversion of hydroquinone to α-arbutin, a powerful skin lightening agent, by amylosucrase

High-yield enzymatic bioconversion of hydroquinone to α-arbutin, a powerful skin lightening agent, by amylosucrase

Amylosucrase: A Versatile Sucrose-Utilizing Transglucosylase for Glycodiversification

Enzymatic modification of daidzin using heterologously expressed amylosucrase in Bacillus subtilis

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