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Pierre Monsan - One of the best experts on this subject based on the ideXlab platform.
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Cloning and Partial Characterization of an Extracellular Dextransucrase Coding Region (DSR-V) from Leuconostoc citreum M-3
2019Co-Authors: Reinaldo Vidal, Magali Remaud-simeon, Sandra Pacios Michelena, Roberto C Aristicas Ribalta, Lisandra Martínez Valdés, Meinardo Lafargue Gámez, Amanda Montes Alvarez, Pierre MonsanAbstract:The Dextransucrase enzymes synthesize dextran, a glucose polymer with broad industrial applications, making the search for new Dextransucrases of great interest. The work described aimed at the partial characterizing of a recombinant Dextransucrase enzyme from Leuconostoc citreum M-3. From the genomic DNA of strain M-3, an amplicon containing a coding region of a Dextransucrase called DSRV was isolated, and deposited in the GenBankTM (Accession number: KF724950). The amino acid sequence alignment of DSR-V with other Dextransucrases demonstrated that it shares a 94% identity with the DSR-D of L. mesenteroides Lcc4 and the DSR-S of L. mesenteroides NRRL B-512F. The DSR-V was cloned and expressed in Escherichia coli JM109 facilitating the formation and detection of DSR-V specific dextran. The SDS-PAGE soluble fraction zymography of E. coli DSR-V and the C-13-NMR spectra of dextran polymers synthesized by this clone confirm that L. citreum M-3 dsrV gene codes for a different Dextransucrase synthesizing linear dextrans with mainly alpha(1-6) linkages.
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A dextran with unique rheological properties produced by the Dextransucrase from Oenococcus kitaharae DSM 17330
Carbohydrate Polymers, 2018Co-Authors: Marlène Vuillemin, Pierre Monsan, Magali Remaud-simeon, Florent Grimaud, Marion Claverie, Agnès Rolland-sabaté, Catherine Garnier, Patrick Lucas, Marguerite Dols-lafargue, Claire MoulisAbstract:A gene encoding a novel Dextransucrase was identified in the genome of Oenococcus kitaharae DSM17330 and cloned into E. coli. With a kcat of 691 s(-1) and a half-life time of 111 h at 30 degrees C, the resulting recombinant enzyme -named DSR-OK- stands as one of the most efficient and stable Dextransucrase characterized to date. From sucrose, this enzyme catalyzes the synthesis of a quasi linear dextran with a molar mass higher than 1 x 10(9) g.mol(-1) that presents uncommon rheological properties such as a higher viscosity than that of the most industrially used dextran from L. mesenteroides NRRL-B-512F, a yield stress that was never described before for any type of dextran, as well as a gel-like structure. All these properties open the way to a vast array of new applications in health, food/feed, bulk or fine chemicals fields.
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a dextran with unique rheological properties produced by the Dextransucrase from oenococcus kitaharae dsm 17330
Carbohydrate Polymers, 2018Co-Authors: Marlène Vuillemin, Magali Remaudsimeon, Pierre Monsan, Florent Grimaud, Marion Claverie, Catherine Garnier, Patrick Lucas, Agnes Rollandsabate, Marguerite Dolslafargue, Claire MoulisAbstract:Abstract A gene encoding a novel Dextransucrase was identified in the genome of Oenococcus kitaharae DSM17330 and cloned into E. coli . With a kcat of 691 s −1 and a half-life time of 111 h at 30 °C, the resulting recombinant enzyme -named DSR-OK- stands as one of the most efficient and stable Dextransucrase characterized to date. From sucrose, this enzyme catalyzes the synthesis of a quasi linear dextran with a molar mass higher than 1 × 10 9 g·mol −1 that presents uncommon rheological properties such as a higher viscosity than that of the most industrially used dextran from L. mesenteroides NRRL-B-512F, a yield stress that was never described before for any type of dextran, as well as a gel-like structure. All these properties open the way to a vast array of new applications in health, food/feed, bulk or fine chemicals fields.
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A novel Dextransucrase is produced by Leuconostoc citreum strain B/110-1-2: an isolate used for the industrial production of dextran and dextran derivatives.
Journal of Industrial Microbiology & Biotechnology, 2011Co-Authors: Reinaldo Fraga Vidal, Sandrine Morel, Magali Remaud-simeon, Claire Moulis, Aidín Martínez, Pierre Escalier, Pierre MonsanAbstract:The industrial Leuconostoc strain B/110-1-2 producing dextran and dextran derivatives was taxonomically identified by 16S rRNA as L. citreum. Its Dextransucrase enzymes were characterized according to their cellular location and reaction specificity. In the presence of sucrose, the strain B/110-1-2 produced two cell-associated Dextransucrases (31.54% of the total glucosyltransferase activity) with molecular weights of 160 and 240 kDa and a soluble Dextransucrase (68.46%) at 160–180 kDa. Two open reading frames (ORF) coding for L. citreum strain B/110-1-2 Dextransucrases were identified. One of them shared a 52% identity with the alternansucrase ASR of L. citreum NRRL B-1355 and with a putative annotated alternansucrase sequence found in the genome of L. citreum KM20. The structural analysis (HPAEC-PAD, HPSEC, and 13C-NMR) of the polymer and oligodextrans produced by the B/110-1-2 Dextransucrases suggest this novel glucansucrase has specificity similar to a Dextransucrase but not to an alternansucrase, producing a soluble linear dextran with glucose molecules linked mainly in α-1,6 and α-1,3 with α-1,4 branches. These results enhance the understanding of this industrially significant strain and will aid in distinguishing between physiologically similar Leuconostoc spp. strains.
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leuconostoc mesenteroides glucansucrase synthesis of flavonoid glucosides by acceptor reactions in aqueous organic solvents
Carbohydrate Research, 2006Co-Authors: Anne Bertrand, Francois Lefoulon, Yves Rolland, Pierre Monsan, Sandrine Morel, Magali RemaudsimeonAbstract:Abstract The enzymatic glucosylation of luteolin was attempted using two glucansucrases: the Dextransucrase from Leuconostoc mesenteroides NRRL B-512F and the alternansucrase from L. mesenteroides NRRL B-23192. Reactions were carried out in aqueous-organic solvents to improve luteolin solubility. A molar conversion of 44% was achieved after 24 h of reaction catalysed by Dextransucrase from L. mesenteroides NRRL B-512F in a mixture of acetate buffer (70%)/bis(2-methoxyethyl) ether (30%). Two products were characterised by nuclear magnetic resonance (NMR) spectroscopy: luteolin-3′-O-α- d -glucopyranoside and luteolin-4′-O-α- d -glucopyranoside. In the presence of alternansucrase from L. mesenteroides NRRL B-23192, three additional products were obtained with a luteolin conversion of 8%. Both enzymes were also able to glucosylate quercetin and myricetin with conversion of 4% and 49%, respectively.
Arun Goyal - One of the best experts on this subject based on the ideXlab platform.
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purification and characterization of Dextransucrase from weissella cibaria rba12 and its application in in vitro synthesis of prebiotic oligosaccharides in mango and pineapple juices
Lwt - Food Science and Technology, 2017Co-Authors: Rwivoo Baruah, Barsha Deka, Arun GoyalAbstract:Abstract Dextransucrase produced by Weissella cibaria RBA12 isolated from pummelo was purified by PEG-400 and PEG-1500 fractionation, followed by gel filtration. The enzyme purified by 0.25 mL/L PEG 400 gave specific activity 410 μkat/g with 25-fold purification. Purified Dextransucrase gave a single, homogeneous protein of molecular size ∼180 kDa on analysis by SDS-PAGE. Purified dexransucrase was optimally active at 40 °C and pH 5.4. It gave maximum velocity (V max ) and Michaelis constant (K m ) of 488.3 μkat/g and 19.2 mmol/L, respectively. The enzyme was thermally stable up to 30 °C and highest pH stability at pH 5.5 for 1 h Mg 2+ and Ca 2+ ions enhanced the enzyme activity by 40% and 25%, respectively. The in-situ production of isomalto-oligosaccharide was carried out by Dextransucrase using mango and pineapple juices. The native sugars (sucrose, glucose and fructose) present in both juices were confirmed by HPLC. The glucose and fructose present in juices acted as acceptor molecules. In both juices, isomalto-oligosaccharides from DP3 to DP5 along with isomaltose (DP2) and leucrose (DP2) were synthesized in situ by Dextransucrase reaction utilizing the native sucrose. Sucrose content of the juices was eliminated resulting in its lower calorific value highlighting the potential of Dextransucrase for production of functional foods.
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optimization of isomaltooligosaccharide size distribution by acceptor reaction of weissella confusa Dextransucrase and characterization of novel α 1 2 branched isomaltooligosaccharides
Journal of Agricultural and Food Chemistry, 2016Co-Authors: Minna Juvonen, Ilkka Kajala, Shraddha Shukla, Hannu Maaheimo, Arun Goyal, Kati Katina, Päivi Tuomainen, Maija TenkanenAbstract:Long-chain isomaltooligosaccharides (IMOs) are promising prebiotics. IMOs were produced by a Weissella confusa Dextransucrase via maltose acceptor reaction. The inputs of substrates (i.e., sucrose and maltose, 0.15–1 M) and Dextransucrase (1–10 U/g sucrose) were used to control IMO yield and profile. According to response surface modeling, 1 M sucrose and 0.5 M maltose were optimal for the synthesis of longer IMOs, whereas the Dextransucrase dosage showed no significant effect. In addition to the principal linear IMOs, a homologous series of minor IMOs were also produced from maltose. As identified by MSn and NMR spectroscopy, the minor trisaccharide contained an α-(1→2)-linked glucosyl residue on the reducing residue of maltose and thus was α-d-glucopyranosyl-(1→2)-[α-d-glucopyranosyl-(1→4)]-d-glucopyranose (centose). The higher members of the series were probably formed by the attachment of a single unit branch to linear IMOs. This is the first report of such α-(1→2)-branched IMOs produced from maltose ...
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optimization of isomaltooligosaccharide size distribution by acceptor reaction of weissella confusa Dextransucrase and characterization of novel α 1 2 branched isomaltooligosaccharides
Journal of Agricultural and Food Chemistry, 2016Co-Authors: Qiao Shi, Minna Juvonen, Ilkka Kajala, Shraddha Shukla, Hannu Maaheimo, Arun Goyal, Kati Katina, Päivi Tuomainen, Yaxi Hou, Maija TenkanenAbstract:Long-chain isomaltooligosaccharides (IMOs) are promising prebiotics. IMOs were produced by a Weissella confusa Dextransucrase via maltose acceptor reaction. The inputs of substrates (i.e., sucrose and maltose, 0.15-1 M) and Dextransucrase (1-10 U/g sucrose) were used to control IMO yield and profile. According to response surface modeling, 1 M sucrose and 0.5 M maltose were optimal for the synthesis of longer IMOs, whereas the Dextransucrase dosage showed no significant effect. In addition to the principal linear IMOs, a homologous series of minor IMOs were also produced from maltose. As identified by MS(n) and NMR spectroscopy, the minor trisaccharide contained an α-(1→2)-linked glucosyl residue on the reducing residue of maltose and thus was α-d-glucopyranosyl-(1→2)-[α-d-glucopyranosyl-(1→4)]-d-glucopyranose (centose). The higher members of the series were probably formed by the attachment of a single unit branch to linear IMOs. This is the first report of such α-(1→2)-branched IMOs produced from maltose by a Dextransucrase.
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structure modeling and functional analysis of recombinant Dextransucrase from weissella confusa cab3 expressed in lactococcus lactis
Preparative Biochemistry & Biotechnology, 2016Co-Authors: Shraddha Shukla, Rwivoo Baruah, Ilkka Kajala, Kati Katina, Maija Tenkanen, Riikka Juvonen, Antti Nyyssölä, Anil Kumar Verma, Arun GoyalAbstract:ABSTRACTThe Dextransucrase gene from Weissella confusa Cab3, having an open reading frame of 4.2 kb coding for 1,402 amino acids, was amplified, cloned, and expressed in Lactococcus lactis. The recombinant Dextransucrase, WcCab3-rDSR was expressed as extracellular enzyme in M17 medium with a specific activity of 1.5 U/mg which after purification by PEG-400 fractionation gave 6.1 U/mg resulting in 4-fold purification. WcCab3-rDSR was expressed as soluble and homogeneous protein of molecular mass, approximately, 180 kDa as analyzed by SDS-PAGE. It displayed maximum enzyme activity at 35°C at pH 5.0 in 50 mM sodium acetate buffer. WcCab3-rDSR gave Km of 6.2 mM and Vm of 6.3 µmol/min/mg. The characterization of dextran synthesized by WcCab3-rDSR by Fourier transform infrared and nuclear magnetic resonance spectroscopic analyses revealed the structural similarities with the dextran produced by the native Dextransucrase. The modeled structure of WcCab3-rDSR using the crystal structures of Dextransucrase from La...
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Optimization of Isomaltooligosaccharide Size Distribution by Acceptor Reaction of Weissella confusa Dextransucrase and Characterization of Novel α‑(1→2)-Branched Isomaltooligosaccharides
2016Co-Authors: Qiao Shi, Minna Juvonen, Ilkka Kajala, Shraddha Shukla, Hannu Maaheimo, Arun Goyal, Kati Katina, Päivi Tuomainen, Yaxi Hou, Maija TenkanenAbstract:Long-chain isomaltooligosaccharides (IMOs) are promising prebiotics. IMOs were produced by a Weissella confusa Dextransucrase via maltose acceptor reaction. The inputs of substrates (i.e., sucrose and maltose, 0.15–1 M) and Dextransucrase (1–10 U/g sucrose) were used to control IMO yield and profile. According to response surface modeling, 1 M sucrose and 0.5 M maltose were optimal for the synthesis of longer IMOs, whereas the Dextransucrase dosage showed no significant effect. In addition to the principal linear IMOs, a homologous series of minor IMOs were also produced from maltose. As identified by MSn and NMR spectroscopy, the minor trisaccharide contained an α-(1→2)-linked glucosyl residue on the reducing residue of maltose and thus was α-d-glucopyranosyl-(1→2)-[α-d-glucopyranosyl-(1→4)]-d-glucopyranose (centose). The higher members of the series were probably formed by the attachment of a single unit branch to linear IMOs. This is the first report of such α-(1→2)-branched IMOs produced from maltose by a Dextransucrase
Maija Tenkanen - One of the best experts on this subject based on the ideXlab platform.
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optimization of isomaltooligosaccharide size distribution by acceptor reaction of weissella confusa Dextransucrase and characterization of novel α 1 2 branched isomaltooligosaccharides
Journal of Agricultural and Food Chemistry, 2016Co-Authors: Minna Juvonen, Ilkka Kajala, Shraddha Shukla, Hannu Maaheimo, Arun Goyal, Kati Katina, Päivi Tuomainen, Maija TenkanenAbstract:Long-chain isomaltooligosaccharides (IMOs) are promising prebiotics. IMOs were produced by a Weissella confusa Dextransucrase via maltose acceptor reaction. The inputs of substrates (i.e., sucrose and maltose, 0.15–1 M) and Dextransucrase (1–10 U/g sucrose) were used to control IMO yield and profile. According to response surface modeling, 1 M sucrose and 0.5 M maltose were optimal for the synthesis of longer IMOs, whereas the Dextransucrase dosage showed no significant effect. In addition to the principal linear IMOs, a homologous series of minor IMOs were also produced from maltose. As identified by MSn and NMR spectroscopy, the minor trisaccharide contained an α-(1→2)-linked glucosyl residue on the reducing residue of maltose and thus was α-d-glucopyranosyl-(1→2)-[α-d-glucopyranosyl-(1→4)]-d-glucopyranose (centose). The higher members of the series were probably formed by the attachment of a single unit branch to linear IMOs. This is the first report of such α-(1→2)-branched IMOs produced from maltose ...
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optimization of isomaltooligosaccharide size distribution by acceptor reaction of weissella confusa Dextransucrase and characterization of novel α 1 2 branched isomaltooligosaccharides
Journal of Agricultural and Food Chemistry, 2016Co-Authors: Qiao Shi, Minna Juvonen, Ilkka Kajala, Shraddha Shukla, Hannu Maaheimo, Arun Goyal, Kati Katina, Päivi Tuomainen, Yaxi Hou, Maija TenkanenAbstract:Long-chain isomaltooligosaccharides (IMOs) are promising prebiotics. IMOs were produced by a Weissella confusa Dextransucrase via maltose acceptor reaction. The inputs of substrates (i.e., sucrose and maltose, 0.15-1 M) and Dextransucrase (1-10 U/g sucrose) were used to control IMO yield and profile. According to response surface modeling, 1 M sucrose and 0.5 M maltose were optimal for the synthesis of longer IMOs, whereas the Dextransucrase dosage showed no significant effect. In addition to the principal linear IMOs, a homologous series of minor IMOs were also produced from maltose. As identified by MS(n) and NMR spectroscopy, the minor trisaccharide contained an α-(1→2)-linked glucosyl residue on the reducing residue of maltose and thus was α-d-glucopyranosyl-(1→2)-[α-d-glucopyranosyl-(1→4)]-d-glucopyranose (centose). The higher members of the series were probably formed by the attachment of a single unit branch to linear IMOs. This is the first report of such α-(1→2)-branched IMOs produced from maltose by a Dextransucrase.
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structure modeling and functional analysis of recombinant Dextransucrase from weissella confusa cab3 expressed in lactococcus lactis
Preparative Biochemistry & Biotechnology, 2016Co-Authors: Shraddha Shukla, Rwivoo Baruah, Ilkka Kajala, Kati Katina, Maija Tenkanen, Riikka Juvonen, Antti Nyyssölä, Anil Kumar Verma, Arun GoyalAbstract:ABSTRACTThe Dextransucrase gene from Weissella confusa Cab3, having an open reading frame of 4.2 kb coding for 1,402 amino acids, was amplified, cloned, and expressed in Lactococcus lactis. The recombinant Dextransucrase, WcCab3-rDSR was expressed as extracellular enzyme in M17 medium with a specific activity of 1.5 U/mg which after purification by PEG-400 fractionation gave 6.1 U/mg resulting in 4-fold purification. WcCab3-rDSR was expressed as soluble and homogeneous protein of molecular mass, approximately, 180 kDa as analyzed by SDS-PAGE. It displayed maximum enzyme activity at 35°C at pH 5.0 in 50 mM sodium acetate buffer. WcCab3-rDSR gave Km of 6.2 mM and Vm of 6.3 µmol/min/mg. The characterization of dextran synthesized by WcCab3-rDSR by Fourier transform infrared and nuclear magnetic resonance spectroscopic analyses revealed the structural similarities with the dextran produced by the native Dextransucrase. The modeled structure of WcCab3-rDSR using the crystal structures of Dextransucrase from La...
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Optimization of Isomaltooligosaccharide Size Distribution by Acceptor Reaction of Weissella confusa Dextransucrase and Characterization of Novel α‑(1→2)-Branched Isomaltooligosaccharides
2016Co-Authors: Qiao Shi, Minna Juvonen, Ilkka Kajala, Shraddha Shukla, Hannu Maaheimo, Arun Goyal, Kati Katina, Päivi Tuomainen, Yaxi Hou, Maija TenkanenAbstract:Long-chain isomaltooligosaccharides (IMOs) are promising prebiotics. IMOs were produced by a Weissella confusa Dextransucrase via maltose acceptor reaction. The inputs of substrates (i.e., sucrose and maltose, 0.15–1 M) and Dextransucrase (1–10 U/g sucrose) were used to control IMO yield and profile. According to response surface modeling, 1 M sucrose and 0.5 M maltose were optimal for the synthesis of longer IMOs, whereas the Dextransucrase dosage showed no significant effect. In addition to the principal linear IMOs, a homologous series of minor IMOs were also produced from maltose. As identified by MSn and NMR spectroscopy, the minor trisaccharide contained an α-(1→2)-linked glucosyl residue on the reducing residue of maltose and thus was α-d-glucopyranosyl-(1→2)-[α-d-glucopyranosyl-(1→4)]-d-glucopyranose (centose). The higher members of the series were probably formed by the attachment of a single unit branch to linear IMOs. This is the first report of such α-(1→2)-branched IMOs produced from maltose by a Dextransucrase
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Structure modeling and functional analysis of recombinant Dextransucrase from Weissella confusa Cab3 expressed in Lactococcus lactis
2016Co-Authors: Shraddha Shukla, Rwivoo Baruah, Ilkka Kajala, Kati Katina, Maija Tenkanen, Riikka Juvonen, Antti Nyyssölä, Anil Kumar Verma, Arun GoyalAbstract:The Dextransucrase gene from Weissella confusa Cab3, having an open reading frame of 4.2 kb coding for 1,402 amino acids, was amplified, cloned, and expressed in Lactococcus lactis. The recombinant Dextransucrase, WcCab3-rDSR was expressed as extracellular enzyme in M17 medium with a specific activity of 1.5 U/mg which after purification by PEG-400 fractionation gave 6.1 U/mg resulting in 4-fold purification. WcCab3-rDSR was expressed as soluble and homogeneous protein of molecular mass, approximately, 180 kDa as analyzed by SDS-PAGE. It displayed maximum enzyme activity at 35°C at pH 5.0 in 50 mM sodium acetate buffer. WcCab3-rDSR gave Km of 6.2 mM and Vm of 6.3 µmol/min/mg. The characterization of dextran synthesized by WcCab3-rDSR by Fourier transform infrared and nuclear magnetic resonance spectroscopic analyses revealed the structural similarities with the dextran produced by the native Dextransucrase. The modeled structure of WcCab3-rDSR using the crystal structures of Dextransucrase from Lactobacillus reuteri (protein data bank, PDB id: 3HZ3) and Streptococcus mutans (PDB id: 3AIB) as templates depicted the presence of different domains such as A, B, C, IV, and V. The domains A and B are circularly permuted in nature having (β/α)8 triose phosphate isomerase-barrel fold making the catalytic core of WcCab3-rDSR. The structure superposition and multiple sequence alignment analyses of WcCab3-rDSR with available structures of enzymes from family 70 GH suggested that the amino acid residue Asp510 acts as a nucleophile, Glu548 acts as a catalytic acid/base, whereas Asp621 acts as a transition-state stabilizer and these residues are found to be conserved within the family.
Claire Moulis - One of the best experts on this subject based on the ideXlab platform.
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A dextran with unique rheological properties produced by the Dextransucrase from Oenococcus kitaharae DSM 17330
Carbohydrate Polymers, 2018Co-Authors: Marlène Vuillemin, Pierre Monsan, Magali Remaud-simeon, Florent Grimaud, Marion Claverie, Agnès Rolland-sabaté, Catherine Garnier, Patrick Lucas, Marguerite Dols-lafargue, Claire MoulisAbstract:A gene encoding a novel Dextransucrase was identified in the genome of Oenococcus kitaharae DSM17330 and cloned into E. coli. With a kcat of 691 s(-1) and a half-life time of 111 h at 30 degrees C, the resulting recombinant enzyme -named DSR-OK- stands as one of the most efficient and stable Dextransucrase characterized to date. From sucrose, this enzyme catalyzes the synthesis of a quasi linear dextran with a molar mass higher than 1 x 10(9) g.mol(-1) that presents uncommon rheological properties such as a higher viscosity than that of the most industrially used dextran from L. mesenteroides NRRL-B-512F, a yield stress that was never described before for any type of dextran, as well as a gel-like structure. All these properties open the way to a vast array of new applications in health, food/feed, bulk or fine chemicals fields.
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a dextran with unique rheological properties produced by the Dextransucrase from oenococcus kitaharae dsm 17330
Carbohydrate Polymers, 2018Co-Authors: Marlène Vuillemin, Magali Remaudsimeon, Pierre Monsan, Florent Grimaud, Marion Claverie, Catherine Garnier, Patrick Lucas, Agnes Rollandsabate, Marguerite Dolslafargue, Claire MoulisAbstract:Abstract A gene encoding a novel Dextransucrase was identified in the genome of Oenococcus kitaharae DSM17330 and cloned into E. coli . With a kcat of 691 s −1 and a half-life time of 111 h at 30 °C, the resulting recombinant enzyme -named DSR-OK- stands as one of the most efficient and stable Dextransucrase characterized to date. From sucrose, this enzyme catalyzes the synthesis of a quasi linear dextran with a molar mass higher than 1 × 10 9 g·mol −1 that presents uncommon rheological properties such as a higher viscosity than that of the most industrially used dextran from L. mesenteroides NRRL-B-512F, a yield stress that was never described before for any type of dextran, as well as a gel-like structure. All these properties open the way to a vast array of new applications in health, food/feed, bulk or fine chemicals fields.
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Overview of the glucansucrase equipment of [i]Leuconostoc citreum[/i] LBAE-E16 and LBAE-C11, two strains isolated from sourdough
FEMS Microbiology Letters, 2014Co-Authors: Myriam Amari, Sandrine Morel, Magali Remaud-simeon, Hervé Robert, Valérie Gabriel, Bruno Gabriel, Claire Moulis, Catherine Fontagné-faucherAbstract:The whole set of putative glucansucrases from Leuconostoc citreum LBAE-E16 and LBAE-C11 was retrieved from the draft genome sequence of these two sourdough strains previously suggested as alternan producers. Four and five putative glycoside hydrolase family 70 (GH70) encoding genes were identified in the genome sequence of strain C11 and E16, respectively. Some putative genes have high sequence identity to known Leuconostoc Dextransucrases. Molecular and biochemical data confirmed that L. citreum C11 could be considered as a new alternan-producing strain, unlike strain E16. In the latter, two new putative glucansucrases with unusual structural features were retrieved. In particular, the GSE16-5 gene encodes for a protein of 2063 amino acids with a theoretical molecular mass of 229 kDa that shares 61% identity with the alternansucrase (ASR) of L. citreum NRRL B-1355, due to the presence of seven APY repeats identified in the C-terminal peptide sequence. Cloning and expression of the corresponding coding sequence revealed synthesis of a low molecular weight (10(4) Da) linear dextran polymer with glucosyl residues only linked by alpha-1,6 linkages. This novel GH70 enzyme may thus be viewed as a natural chimeric enzyme resulting from the addition of the ASR C-terminal region in a Dextransucrase.
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Characterization of a novel Dextransucrase from Weissella confusa isolated from sourdough
Applied Microbiology and Biotechnology, 2013Co-Authors: Myriam Amari, Sandrine Morel, Magali Remaud-simeon, Hervé Robert, Valérie Gabriel, Bruno Gabriel, Luisa Fernanda Gomez Arango, Claire Moulis, Catherine Fontagné-faucherAbstract:Weissella confusa and Weissella cibaria isolated from wheat sourdoughs produce, from sucrose, linear dextrans due to a single soluble Dextransucrase. In this study, the first complete gene sequence encoding Dextransucrase from a W. confusa strain (LBAE C39-2) along with the one from a W. cibaria strain (LBAE K39) were reported. Corresponding gene cloning was achieved using specific primers designed on the basis of the draft genome sequence of these species. Deduced amino acid sequence of W. confusa and W. cibaria Dextransucrase revealed common structural features of the glycoside hydrolase family 70. Notably, the regions located in the vicinity of the catalytic triad (D, E, D) are highly conserved. However, comparison analysis also revealed that Weissella Dextransucrases form a distinct phylogenetic group within glucansucrases of other lactic acid bacteria. We then cloned the W. confusa C39-2 Dextransucrase gene and successfully expressed the mature corresponding enzyme in Escherichia coli. The purified recombinant enzyme rDSRC39-2 catalyzed dextran synthesis from sucrose with a K (m) of 8.6 mM and a V (max) of 20 mu mol/mg/min. According to H-1 and C-13 NMR analysis, the polymer is a linear class 1 dextran with 97.2 % alpha-(1 -> 6) linkages and 2.8 % alpha-(1 -> 3) branch linkages, similar to the one produced by W. confusa C39-2 strain. The enzyme exhibited optimum catalytic activity for temperatures ranging from 35 to 40 A degrees C and a pH of 5.4 in 20 mM sodium acetate buffer. This novel Dextransucrase is responsible for production of dextran with predominant alpha-(1 -> 6) linkages that could find applications as food hydrocolloids.
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A novel Dextransucrase is produced by Leuconostoc citreum strain B/110-1-2: an isolate used for the industrial production of dextran and dextran derivatives.
Journal of Industrial Microbiology & Biotechnology, 2011Co-Authors: Reinaldo Fraga Vidal, Sandrine Morel, Magali Remaud-simeon, Claire Moulis, Aidín Martínez, Pierre Escalier, Pierre MonsanAbstract:The industrial Leuconostoc strain B/110-1-2 producing dextran and dextran derivatives was taxonomically identified by 16S rRNA as L. citreum. Its Dextransucrase enzymes were characterized according to their cellular location and reaction specificity. In the presence of sucrose, the strain B/110-1-2 produced two cell-associated Dextransucrases (31.54% of the total glucosyltransferase activity) with molecular weights of 160 and 240 kDa and a soluble Dextransucrase (68.46%) at 160–180 kDa. Two open reading frames (ORF) coding for L. citreum strain B/110-1-2 Dextransucrases were identified. One of them shared a 52% identity with the alternansucrase ASR of L. citreum NRRL B-1355 and with a putative annotated alternansucrase sequence found in the genome of L. citreum KM20. The structural analysis (HPAEC-PAD, HPSEC, and 13C-NMR) of the polymer and oligodextrans produced by the B/110-1-2 Dextransucrases suggest this novel glucansucrase has specificity similar to a Dextransucrase but not to an alternansucrase, producing a soluble linear dextran with glucose molecules linked mainly in α-1,6 and α-1,3 with α-1,4 branches. These results enhance the understanding of this industrially significant strain and will aid in distinguishing between physiologically similar Leuconostoc spp. strains.
Doman Kim - One of the best experts on this subject based on the ideXlab platform.
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synthesis and functional characterization of caffeic acid glucoside using leuconostoc mesenteroides Dextransucrase
Journal of Agricultural and Food Chemistry, 2017Co-Authors: Seunghee Nam, Youngmin Kim, Marie K Walsh, Youngjung Wee, Kwangyeol Yang, Songhee Han, Thi Thanh Hanh Nguyen, Ji Young Kim, Doman KimAbstract:Caffeic acid was modified via transglucosylation using sucrose and Dextransucrase from Leuconostoc mesenteroides B-512FMCM. Following enzymatic modification, a caffeic acid glucoside was isolated by butanol separation, silica gel chromatography, and preparative HPLC. The synthesized caffeic acid glucoside had a molecular mass-to-charge ratio of 365 m/z, and its structure was identified as caffeic acid-3-O-α-d-glucopyranoside. The production of this caffeic acid-3-O-α-d-glucopyranoside at a concentration of 153 mM was optimized using 325 mM caffeic acid, 355 mM sucrose, and 650 mU mL–1 Dextransucrase in the synthesis reaction. In comparison with the caffeic acid, the caffeic acid-3-O-α-d-glucopyranoside displayed 3-fold higher water solubility, 1.66-fold higher antilipid peroxidation effect, 15% stronger inhibition of colon cancer cell growth, and 11.5-fold higher browning resistance. These results indicate that this caffeic acid-3-O-α-d-glucopyranoside may be a suitable functional component of food and ph...
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Transglycosylation of gallic acid by using Leuconostoc glucansucrase and its characterization as a functional cosmetic agent
SpringerOpen, 2017Co-Authors: Seunghee Nam, Doman Kim, Jeongjin Park, Woojin Jun, A. Abd M. El-aty, Jin Young Choi, Do-ik Kim, Kwangyeol YangAbstract:Abstract Gallic acid glycoside was enzymatically synthesized by using Dextransucrase and sucrose from gallic acid. After purification by butanol partitioning and preparative HPLC, gallic acid glucoside was detected at m/z 355 (C13, H16, O10, Na)+ by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The yield of gallic acid glucoside was found to be 35.7% (114 mM) by response surface methodology using a reaction mixture of 319 mM gallic acid, 355 mM sucrose, and 930 mU/mL Dextransucrase. The gallic acid glucoside obtained showed 31% higher anti-lipid peroxidation and stronger inhibition (Ki = 1.23 mM) against tyrosinase than that shown by gallic acid (Ki = 1.98 mM). In UVB-irradiated human fibroblast cells, gallic acid glucoside lowered matrix metalloproteinase-1 levels and increased the collagen content, which was indicative of a stronger anti-aging effect than that of gallic acid or arbutin. These results indicated that gallic acid glucoside is likely a superior cosmetic ingredient with skin-whitening and anti-aging functions
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Synthesis and Functional Characterization of Caffeic Acid Glucoside Using Leuconostoc mesenteroides Dextransucrase
2017Co-Authors: Seunghee Nam, Youngmin Kim, Marie K Walsh, Youngjung Wee, Kwangyeol Yang, Songhee Han, Thi Thanh Hanh Nguyen, Ji Young Kim, Doman KimAbstract:Caffeic acid was modified via transglucosylation using sucrose and Dextransucrase from Leuconostoc mesenteroides B-512FMCM. Following enzymatic modification, a caffeic acid glucoside was isolated by butanol separation, silica gel chromatography, and preparative HPLC. The synthesized caffeic acid glucoside had a molecular mass-to-charge ratio of 365 m/z, and its structure was identified as caffeic acid-3-O-α-d-glucopyranoside. The production of this caffeic acid-3-O-α-d-glucopyranoside at a concentration of 153 mM was optimized using 325 mM caffeic acid, 355 mM sucrose, and 650 mU mL–1 Dextransucrase in the synthesis reaction. In comparison with the caffeic acid, the caffeic acid-3-O-α-d-glucopyranoside displayed 3-fold higher water solubility, 1.66-fold higher antilipid peroxidation effect, 15% stronger inhibition of colon cancer cell growth, and 11.5-fold higher browning resistance. These results indicate that this caffeic acid-3-O-α-d-glucopyranoside may be a suitable functional component of food and pharmaceutical products
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construction of a fusion enzyme of Dextransucrase and dextranase application for one step synthesis of isomalto oligosaccharides
Enzyme and Microbial Technology, 2009Co-Authors: Youngmin Kim, Hee-kyoung Kang, Miyoung Seo, Kimura Atsuo, Doman KimAbstract:Abstract The linear isomalto-oligosaccharides (IMO) with DP2–DP10 were produced by one-step process using engineered fusion enzyme (DXSR) of endo-dextranase and only α-(1–6) glucan synthesizing Dextransucrase. The fusion enzyme was successfully expressed in Escherichia coli and characterized. Compared to individual enzymes, DXSR had 150% increased endo-dextranase activity and 98% decreased Dextransucrase activity. The partially purified DXSR displayed molecular mass of 240 kDa as analyzed by SDS–PAGE. It showed both enzyme activities on analysis by zymogram. The thermal- and pH-stability of DXSR was around 28 °C and pH at 5.0–6.4, respectively. IMOs production by DXSR was increased by the addition of metal ions such as Fe 2+ , Li + , K + and Ni 2+ , but the enzyme was strongly inhibited by Hg 2+ and Ag + . DXSR produced linear IMO with DP2–DP10 using sucrose as a sole substrate. The molecular weight and amount of IMO could be controlled by the sucrose concentration. DXSR gave 30-fold higher production of IMO than that of an equal activity mixture of the two enzymes such as dextranase and Dextransucrase.
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directed evolution of a Dextransucrase for increased constitutive activity and the synthesis of a highly branched dextran
Journal of Molecular Catalysis B-enzymatic, 2003Co-Authors: Hee-kyoung Kang, John F Robyt, Eunseong Seo, Doman KimAbstract:Abstract An Escherichia coli transformant (pDSRB742CK) was obtained from the DSRB742 clone by using ultrasoft X-rays for the expression of a Dextransucrase. The enzyme differed in several aspects from DSRB742 Dextransucrase: it (1) was constitutive; (2) was extracellular; (3) had 2.6 times greater activity (0.035 IU/ml and 0.23 IU/mg); and (4) synthesized a highly (15.6%) α-(1→3) branched dextran. Seven nucleotides of the parent gene (dsrB742) were changed in the nucleotide sequence; four nucleotides were changed in the open reading frame (ORF) that resulted in a 30 amino acid deletion in the N-terminus.