390 results on '"Amylosucrase"'
Search Results
352. Fluorescence detection of the transglycosylation activity of amylosucrase.
- Author
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Seo DH, Jung JH, and Park CS
- Subjects
- Chromatography, High Pressure Liquid, Fluorescence, Glycosylation, Kinetics, Bacterial Proteins metabolism, Deinococcus enzymology, Glucosyltransferases metabolism, Neisseria enzymology, Sucrose metabolism
- Abstract
The purpose of this study was to investigate the novel fluorescence-based assay for the transglycosylation activity of amylosucrase (ASase). The transglycosylation activity of ASase from Deinococcus geothermalis (DGAS), ASase from Neisseria polysaccharea (NPAS), and DGAS-B (chimeric ASase wherein the B domain from DGAS was exchanged with the B domain of NPAS in a DGAS background) was applied to modify 4-methlylumberlliferone (MU) to 4-methylumberlliferone glucoside (MUG) using MU as an acceptor and sucrose as a glucoside donor. The result of HPLC (high performance liquid chromatography) show that the bioconversion of MUG with ASases was successfully accomplished using sucrose and MU. Kinetic studies of ASases were performed to determine kinetic parameter for sucrose and MU. The order of overall performance (k
cat /Km ) of transglycosylation activity for MU among DGAS, DGAS-B and NPAS was as follows: DGAS-B (8.1) > DGAS (5.0) > NPAS (0.4). The fluorescence-based transglycosylation assay using MU has a potential to be used as the detection of transglycosylation activity of ASase and to screen novel ASase variants, which may be improved in their transglycosylation activities., (Copyright © 2017 Elsevier Inc. All rights reserved.)- Published
- 2017
- Full Text
- View/download PDF
353. Preparation and characterization of the inclusion complexes between amylosucrase-treated waxy starch and palmitic acid.
- Author
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Kim HI, Kim HR, Choi SJ, Park CS, and Moon TW
- Abstract
Amylosucrase-treated waxy corn starch (AS) was produced to extend the chain length of amylopectin to a great extent in comparison to its native chain length. An amylopectin-palmitic acid (PA) complex was prepared by heat-treating (121°C) a starch/PA mixture and its subsequent further incubation (95°C, 24 h); moreover, its structure and digestibility were studied. Unmodified waxy starch could not complex at all, whereas elongation due to amylosucrase modification allowed amylopectin to form a complex with PA to a small extent. Complexation between AS and PA caused a decrease in relative crystallinity. The AS-PA complex displayed an endothermic peak representing type I inclusion complexes rather than type II complexes. The formation of complexes did not significantly affect the in vitro digestibility maintaining the low digestibility of AS resulting from extremely small amounts of complexes and the type of complex.
- Published
- 2017
- Full Text
- View/download PDF
354. The structural characteristics of amylosucrase-treated waxy corn starch and relationship between its in vitro digestibility.
- Author
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Park CS and Park I
- Abstract
The glucotransferase amylosucrase (AS) influences the structural properties of starch, but its precise effects are unclear. The structural characteristics and in vitro digestibility of waxy corn starch modified by AS from Neisseria polysaccharea were examined. AS-treated starch exhibited a higher slowly digestible starch (SDS) fraction, the weak B-type polymorph, lower relative crystallinity, and lower double helix content than those of native starches based on X-ray diffractometry, solid-state
13 C CP/MAS NMR, and FT-IR. AS-treated starches exhibited increased proportions of degree of polymerization (DP) 25-36 and DP≥37 chains. Higher SDS and resistant (RS) fractions, higher proportions of DP 25-36 and DP≥37 chains, more double helices, higher relative crystallinity, and less difference between double helix and relative crystallinity were observed for starch treated with 460 U than with 230 U of AS. AS re-built the double-helical and rearranged crystalline structure of gelatinized starch and consequently influenced the SDS and RS fractions.- Published
- 2017
- Full Text
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355. Impact of amylosucrase modification on the structural and physicochemical properties of native and acid-thinned waxy corn starch.
- Author
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Zhang H, Zhou X, He J, Wang T, Luo X, Wang L, Wang R, and Chen Z
- Subjects
- Hydrolysis, Molecular Structure, Temperature, Thermodynamics, Viscosity, X-Ray Diffraction, Amylopectin chemistry, Glucosyltransferases chemistry, Neisseria enzymology, Starch chemistry, Zea mays chemistry
- Abstract
Recombinant amylosucrase from Neisseria polysaccharea was utilized to modify native and acid-thinned starches. The molecular structures and physicochemical properties of modified starches were investigated. Acid-thinned starch displayed much lower viscosity after gelatinization than did the native starch. However, the enzyme exhibited similar catalytic efficiency for both forms of starch. The modified starches had higher proportions of long (DP>33) and intermediate chains (DP 13-33), and X-ray diffraction showed a B-type crystalline structure for all modified starches. With increasing reaction time, the relative crystallinity and endothermic enthalpy of the modified starches gradually decreased, whereas the melting peak temperatures and resistant starch contents increased. Slight differences were observed in thermal parameters, relative crystallinity, and branch chain length distribution between the modified native and acid-thinned starches. Moreover, the digestibility of the modified starches was not affected by acid hydrolysis pretreatment, but was affected by the percentage of intermediate and long chains., (Copyright © 2016 Elsevier Ltd. All rights reserved.)
- Published
- 2017
- Full Text
- View/download PDF
356. Novel product specificity toward erlose and panose exhibited by multisite engineered mutants of amylosucrase.
- Author
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Vergès A, Cambon E, Barbe S, Moulis C, Remaud-Siméon M, and André I
- Subjects
- Mutation, Missense, Substrate Specificity genetics, Amino Acid Substitution, Bacterial Proteins chemistry, Bacterial Proteins genetics, Glucans chemistry, Glucosyltransferases chemistry, Glucosyltransferases genetics, Neisseria enzymology, Neisseria genetics, Trisaccharides chemistry
- Abstract
A computer-aided engineering approach recently enabled to deeply reshape the active site of N. polysaccharea amylosucrase for recognition of non-natural acceptor substrates. Libraries of variants were constructed and screened on sucrose allowing the identification of 17 mutants able to synthesize molecules from sole sucrose, which are not synthesized by the parental wild-type enzyme. Three of the isolated mutants as well as the new products synthesized were characterized in details. Mutants contain between 7 and 11 mutations in the active site and the new molecules were identified as being a sucrose derivative, named erlose (α-d-glucopyranosyl-(1→4)-α-d-glucopyranosyl-(1→2)-β-d-Fructose), and a new malto-oligosaccharide named panose (α-d-glucopyranosyl-(1→6)-α-d-glucopyranosyl-(1→4)-α-d-Glucose). These product specificities were never reported for none of the amylosucrases characterized to date, nor their engineered variants. Optimization of the production of these trisaccharides of potential interest as sweeteners or prebiotic molecules was carried out. Molecular modelling studies were also performed to shed some light on the molecular factors involved in the novel product specificities of these amylosucrase variants., (© 2016 The Protein Society.)
- Published
- 2017
- Full Text
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357. Studies on a recombinant amylosucrase
- Author
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I. Varlet, F. Albaret, B. Canard, Magali Remaud-Simeon, P. Colonna, Pierre Monsan, and R.M. Willemot
- Subjects
chemistry.chemical_classification ,Amylosucrase activity ,Lysis ,Glycogen ,Biology ,Molecular biology ,law.invention ,chemistry.chemical_compound ,Amylosucrase ,Enzyme ,Biochemistry ,chemistry ,law ,Recombinant DNA ,biology.protein ,Specific activity ,DNA - Abstract
In order to characterize a recombinant amylosucrase activity (E.C. 2.4.1.4.) and to evaluate its potential use as a glucosylation tool, chromosomal Sau 3A DNA fragments from Neisseria polysaccharea were cloned into the phage λ EMBL3. A recombinant phage expressing the amylosucrase activity was isolated. Production of the enzyme was carried out by infection of liquid culture of E . coli . The enzyme was purified from culture lysate to a specific activity of 0.3 U/mg. When incubated with sucrose and traces of glycogen, the recombinant amylosucrase produced an insoluble glucopolysaccharide mainly composed of α-(1 → 4) glucosidic linkages and a very low degree of α-(1 → 6) branched linkages (less than 5 %). The recombinant enzyme is activated by glycogen, starch and maltooligosaccharides. It also catalyzes the transfer of glucosyl residue from sucrose onto a maltopentaose acceptor to produce maltohexaose and heptaose.
- Published
- 1995
358. The effect of delmopinol on glucosyltransferase adsorbed on to saliva-coated hydroxyapatite
- Author
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D. Beeman, William H. Bowen, and D. Steinberg
- Subjects
Saliva ,Morpholines ,Streptococcus mutans ,Amylosucrase ,Surface-Active Agents ,Adsorption ,stomatognathic system ,Desorption ,Humans ,General Dentistry ,Glucans ,chemistry.chemical_classification ,biology ,Chemistry ,Sodium Dodecyl Sulfate ,Biological activity ,Cell Biology ,General Medicine ,biology.organism_classification ,Solutions ,stomatognathic diseases ,Enzyme ,Durapatite ,Otorhinolaryngology ,Biochemistry ,Glucosyltransferases ,biology.protein ,Glucosyltransferase ,Electrophoresis, Polyacrylamide Gel ,Hydroxyapatites ,Nuclear chemistry - Abstract
The aim was to explore the effects of delmopinol, a substituted amino-alcohol compound recently reported as a potential antiplaque agent, on GTF activity in solution and when adsorbed on to sHA. Delmopinol was without a significant effect on GTF activity in solution. In contrast, a reduction in the bound glucans synthesized by the adsorbed GTF was found in the presence of delmopinol. Delmopinol did not displace the adsorbed GTF from the sHA, nor was there significant desorption of glucans from sHA. The total glucan synthesis (bound and unbound) was reduced in the presence of delmopinol. Inhibition of GTF was not reversed by sucrose. Inhibition of GTF activity by delmopinol apparently results from drug-enzyme interaction on the surface of sHA beads. These observations provide further support for the important differences in the properties of adsorbed GTF and GTF in solution, illustrating that GTF-drug interaction differs between enzyme adsorbed to surfaces and enzyme in solution.
- Published
- 1992
359. Cloning of amylosucrase from Alteromonas macleodii and expression using pGEX-4T-1 vector
- Author
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Gil-Yong Lee, Cheon-Seok Park, Dong-Ho Seo, Jong-Hyun Jung, and Suk-Jin Ha
- Subjects
Genetics ,Cloning ,Amylosucrase ,biology.protein ,Bioengineering ,General Medicine ,Vector (molecular biology) ,Biology ,Alteromonas macleodii ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Biotechnology - Published
- 2008
360. Self-Association and Crystallization of Amylose
- Author
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Alain Buléon, Jean-Luc Putaux, Gabrielle Véronèse, Unité de recherche sur les Biopolymères, Interactions Assemblages (BIA), Institut National de la Recherche Agronomique (INRA), Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés (LISBP), Institut National de la Recherche Agronomique (INRA)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS), Centre de Recherches sur les Macromolécules Végétales (CERMAV), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), Centre National de la Recherche Scientifique (CNRS)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Recherche Agronomique (INRA), Centre National de la Recherche Scientifique (CNRS), Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), and Université Joseph Fourier - Grenoble 1 (UJF)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)
- Subjects
[SDV.SA]Life Sciences [q-bio]/Agricultural sciences ,Recrystallization (geology) ,MOLECULAR STRUCTURE ,Starch ,02 engineering and technology ,010402 general chemistry ,01 natural sciences ,law.invention ,chemistry.chemical_compound ,Amylosucrase ,law ,Amylose ,SELF ASSOCIATION ,BIOSYNTHESIS ,IN VITRO SYNTHESIS ,Crystallization ,SYNTHESE ,chemistry.chemical_classification ,biology ,STARCH GRANULE ,Chemistry ,food and beverages ,General Chemistry ,Polymer ,Carbohydrate ,021001 nanoscience & nanotechnology ,0104 chemical sciences ,Crystallography ,Monomer ,Chemical engineering ,biology.protein ,CRYSTALLIZATION ,0210 nano-technology - Abstract
International audience; Amylose, the linear constituent of starch, consists of a(1,4)-linked glucose monomers. Although weakly involved in the crystalline structure of starch, it can be recrystallized in a variety of allomorphic types, including those encountered in native starch (A- and B-types). Amylose can either be extracted from starch or produced in vitro by enzymatic synthesis using amylosucrase or phosphorylase. Recrystallization and self-association of amylose in aqueous solutions have been widely studied to understand both the crystallization of starch during biosynthesis and the structural changes that occur during starch processing. Depending on the chain length, concentration, and temperature, gels, spherulites, or lamellar crystals can be formed withA or B allomorphic type. Other ligand-dependent allomorphs (the various V-types) are obtained when amylose is complexed with molecules such as alcohols, lipids, or flavours. Amylose also self-associates into networks, spherulites, or axialites during in-vitro enzymatic synthesis by amylosucrase. When a highly branched acceptor like glycogen is used, dendritic nanoparticles are formed by elongation of the external chains. The recrystallization of amylose extracted from starch and the self-association of amylose during its in-vitro synthesis are described. The amylose properties are discussed in terms of polymer behaviour and model systems to investigate the structure and formation of starch granules.
- Published
- 2007
361. Effect of short-chain fatty acids on the formation of amylose microparticles by amylosucrase.
- Author
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Lim MC, Park KH, Choi JH, Lee DH, Letona CAM, Baik MY, Park CS, and Kim YR
- Subjects
- Deinococcus enzymology, Models, Molecular, Molecular Conformation, Amylose chemistry, Fatty Acids, Volatile chemistry, Glucosyltransferases metabolism, Microspheres
- Abstract
Amylose microparticles can be produced by self-assembly of amylose molecules through an amylosucrase-mediated synthesis. Here we investigated the role of short-chain fatty acids in the formation of amylose microparticles and the fate of these fatty acids at the end of the reaction. The rate of self-assembly and production yields of amylose microparticles were significantly enhanced in the presence of fatty acids. The effect was dependent on the length of the fatty acid carbon tail; butanoic acid (C4) was the most effective, followed by hexanoic acid (C6) and octanoic acid (C8). The amylose microparticles were investigated by carrying out SEM, XRD, Raman, NMR, FT-IR and DSC analysis. The size, morphology and crystal structure of the resulting amylose microparticles were comparable with those of amylose microparticles produced without fatty acids. The results indicated the carboxyl group of the fatty acid to be responsible for promoting the self-assembly of amylose chains to form microparticles. The fatty acids were eventually removed from the microstructure through the tight association of amylose double helices to form the amylose microparticles., (Copyright © 2016 Elsevier Ltd. All rights reserved.)
- Published
- 2016
- Full Text
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362. Flavanone and isoflavone glucosylation by non-Leloir glycosyltransferases.
- Author
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Overwin H, Wray V, Seeger M, Sepúlveda-Boza S, and Hofer B
- Subjects
- Bacterial Proteins genetics, Escherichia coli genetics, Escherichia coli metabolism, Flavanones chemistry, Glucosyltransferases genetics, Glycosylation, Glycosyltransferases genetics, Isoflavones chemistry, Recombinant Proteins genetics, Bacterial Proteins metabolism, Flavanones metabolism, Glucosyltransferases metabolism, Glycosyltransferases metabolism, Isoflavones metabolism, Recombinant Proteins metabolism
- Abstract
Flavonoids possess a wide range of biological activities. Their glycosylation is of considerable interest, as it often positively influences their pharmacokinetic and other molecular properties. We recently showed that two non-Leloir glycosyltransferases that use sucrose as carbohydrate donor, an amylosucrase from Neisseria polysaccharea (Ams-Np) and a glucansucrase from Streptococcus oralis (GtfR-So), were hardly able to glucosylate flavones, but accepted flavanes as substrates. We now examined compounds from two other flavonoid classes, flavanones and isoflavones for glucose transfer by these enzymes. Taxifolin was investigated as a flavanone analogue of both, the accepted pentahydroxyflavane catechin and the non-accepted pentahydroxyflavone quercetin. It was glucosylated by both enzymes, but much better by GtfR-So than by Ams-Np due to apparent strong inhibition of the latter. The acceptance of a collection of isoflavones strongly depended on the substitution pattern of the core. Only two of the 10 compounds examined yielded glucosides in satisfactory amounts. With these substrates, both enzymes catalyzed formation of a range of products, differing in the number of saccharide units. The structures of mono- and diglycosylated compounds obtained in higher amounts were elucidated. While GtfR-So attached glucose to taxifolin in the B ring at O4', both enzymes glucosylated the isoflavones in the A ring at O7. All products were α-glucosides. Interglycosidic linkages formed by Ams-Np were α1-4. To our knowledge, this is the first report of glucosylation of flavanone and isoflavone aglycones by an amylosucrase. All characterized compounds have not previously been described., (Copyright © 2016 Elsevier B.V. All rights reserved.)
- Published
- 2016
- Full Text
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363. GH13 amylosucrases and GH70 branching sucrases, atypical enzymes in their respective families.
- Author
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Moulis C, André I, and Remaud-Simeon M
- Subjects
- Amino Acid Sequence, Glucosyltransferases chemistry, Glycosyltransferases chemistry, Kinetics, Models, Molecular, Protein Engineering, Glucosyltransferases metabolism, Glycosyltransferases metabolism, Multigene Family
- Abstract
Amylosucrases and branching sucrases are α-retaining transglucosylases found in the glycoside-hydrolase families 13 and 70, respectively, of the clan GH-H. These enzymes display unique activities in their respective families. Using sucrose as substrate and without mediation of nucleotide-activated sugars, amylosucrase catalyzes the formation of an α-(1 → 4) linked glucan that resembles amylose. In contrast, the recently discovered branching sucrases are unable to catalyze polymerization of glucosyl units as they are rather specific for dextran branching through α-(1 → 2) or α-(1 → 3) branching linkages depending on the enzyme regiospecificity. In addition, GH13 amylosucrases and GH70 branching sucrases are naturally promiscuous and can glucosylate different types of acceptor molecules including sugars, polyols, or flavonoids. Amylosucrases have been the most investigated glucansucrases, in particular to control product profiles or to successfully develop tailored α-transglucosylases able to glucosylate various molecules of interest, for example, chemically protected carbohydrates that are planned to enter in chemoenzymatic pathways. The structural traits of these atypical enzymes will be described and compared, and an overview of the potential of natural or engineered enzymes for glycodiversification and chemoenzymatic synthesis will be highlighted.
- Published
- 2016
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364. New insights into the action mode of amylosucrase on amylopectin.
- Author
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Zhang H, Zhou X, Wang T, Luo X, Wang L, Li Y, Wang R, and Chen Z
- Subjects
- Sucrose chemistry, Surface Properties, Waxes chemistry, Zea mays chemistry, Amylopectin chemistry, Glucosyltransferases chemistry, Starch chemistry
- Abstract
To investigate the action mode of amylosucrase (AS) on amylopectin, waxy corn starch (WCS) was selected as an acceptor. The effects of WCS dissolution method, reaction temperature, sucrose concentration and AS activity on transglycosylation degree (TD) were investigated. Under the selected reaction condition, the enzymatic reaction process was divided into two stages, i.e. before and after 0.25h, of which the relations between TD value and reaction time were successfully described using a linear and a logarithmic function, respectively. Then, the elongated WCSs with different TDs were produced according to the theoretical reaction time calculated based on the regression equations. The chain length distribution of the elongated WCSs indicated that all of the branch chains of WCS were greatly elongated by AS before occurrence of starch precipitation. Afterwards, however, AS merely elongated the branch chains whose non-reducing ends were exposed on the surface of the precipitate., (Copyright © 2016 Elsevier B.V. All rights reserved.)
- Published
- 2016
- Full Text
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365. Production of an in Vitro Low-Digestible Starch via Hydrothermal Treatment of Amylosucrase-Modified Normal and Waxy Rice Starches and Its Structural Properties.
- Author
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Kim JH, Kim HR, Choi SJ, Park CS, and Moon TW
- Subjects
- Biocatalysis, Digestion, Hot Temperature, Humans, Models, Biological, Oryza metabolism, Starch metabolism, Viscosity, Glucosyltransferases chemistry, Oryza chemistry, Starch chemistry
- Abstract
We investigated dual modification of normal and waxy rice starch, focusing on digestibility. Amylosucrase (AS) was applied to maximize the slowly digestible and resistant starch fractions. AS-modified starches were adjusted to 25-40% moisture levels and heated at 100 °C for 40 min. AS-modified starches exhibited a B-type crystalline structure, and hydrothermal treatment (HTT) significantly (p < 0.05) increased the relative crystallinity with moisture level. The thermal transition properties of modified starches were also affected by the moisture level. The contents of rapidly digestible starch fraction in AS-modified normal and waxy starches (43.3 ± 3.9 and 18.1 ± 0.6%) decreased to 13.0 ± 1.0 and 0.3 ± 0.3% after HTT, accordingly increasing the low digestible fractions. Although the strengthened crystalline structures of AS-modified starches by HTT were not stable enough to maintain their rigidity under cooking, application of AS and HTT was more effective in waxy rice starch than normal rice starch when lowering digestibility.
- Published
- 2016
- Full Text
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366. Enzymatic Process for High-Yield Turanose Production and Its Potential Property as an Adipogenesis Regulator.
- Author
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Park MO, Lee BH, Lim E, Lim JY, Kim Y, Park CS, Lee HG, Kang HK, and Yoo SH
- Subjects
- 3T3-L1 Cells drug effects, 3T3-L1 Cells metabolism, Animals, Biotechnology methods, Disaccharides metabolism, Fructose metabolism, Gene Expression Regulation drug effects, Glucosyltransferases chemistry, Lipid Metabolism drug effects, Mice, PPAR gamma genetics, Sterol Regulatory Element Binding Protein 1 genetics, Sucrose metabolism, fas Receptor genetics, Adipogenesis drug effects, Disaccharides biosynthesis, Disaccharides pharmacology, Glucosyltransferases metabolism
- Abstract
Turanose is a sucrose isomer naturally existing in honey and a promising functional sweetener due to its low glycemic response. In this study, the extrinsic fructose effect on turanose productivity was examined in Neisseria amylosucrase reaction. Turanose was produced, by increasing the amount of extrinsic fructose as a reaction modulator, with high concentration of sucrose substrate, which resulted in 73.7% of production yield. In physiological functionality test, lipid accumulation in 3T3-L1 preadipocytes in the presence of high amounts of pure glucose was attenuated by turanose substitution in a dose-dependent manner. Turanose treatments at concentrations representing 50%, 75%, and 100% of total glucose concentration in cell media significantly reduced lipid accumulation by 18%, 35%, and 72%, respectively, as compared to controls. This result suggested that turanose had a positive role in controlling adipogenesis, and enzymatic process of turanose production has a potential to develop a functional food ingredient for controlling obesity and related chronic diseases.
- Published
- 2016
- Full Text
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367. Preparation of slowly digestible sweet potato Daeyumi starch by dual enzyme modification.
- Author
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Jo AR, Kim HR, Choi SJ, Lee JS, Chung MN, Han SK, Park CS, and Moon TW
- Subjects
- Amylopectin chemistry, Amylopectin isolation & purification, Animals, Digestion, Neisseria, Pancreatin metabolism, Streptococcus mutans, Swine, Temperature, Thermodynamics, 1,4-alpha-Glucan Branching Enzyme metabolism, Amylopectin metabolism, Glucosyltransferases metabolism, Ipomoea batatas chemistry
- Abstract
Sweet potato Daeyumi starch was dually modified using glycogen branching enzyme (BE) from Streptococcus mutans and amylosucrase (AS) from Neisseria polysaccharea to prepare slowly digestible starch (SDS). Dually modified starches had higher SDS and resistant starch (RS) contents than control starch. The branched chain length distributions of the BE-modified starches indicated an increase in short side-chains [degree of polymerization (DP)≤12] compared with native starch. AS treatment of the BE-modified starches decreased the proportion of short side-chains and increased the proportion of long side-chains (DP≥25) and molecular mass. It also resulted in a B-type X-ray diffraction pattern and an increased relative crystallinity. Regarding thermal properties, the BE-modified starches showed no endothermic peak, whereas the BEAS-modified starches had a broader melting temperature range and lower melting enthalpy compared to native starch. The combined enzymatic treatment resulted in novel glucan polymers with slow digestion properties., (Copyright © 2016 Elsevier Ltd. All rights reserved.)
- Published
- 2016
- Full Text
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368. Efficient Biocatalytic Production of Cyclodextrins by Combined Action of Amylosucrase and Cyclodextrin Glucanotransferase.
- Author
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Koh DW, Park MO, Choi SW, Lee BH, and Yoo SH
- Subjects
- Bacillus enzymology, Bacterial Proteins metabolism, Biocatalysis, Cyclodextrins metabolism, Glucosyltransferases metabolism, Starch chemistry, Starch metabolism, Bacterial Proteins chemistry, Cyclodextrins chemistry, Glucosyltransferases chemistry, Neisseria enzymology
- Abstract
A novel enzymatic process for cyclodextrin (CD) production was developed by utilizing sucrose as raw material instead of corn starch. Cyclodextrin glucanotransferase (CGTase) from Bacillus macerans was applied to produce the CDs from linear α-(1,4)-glucans, which were obtained by Neisseria polysaccharea amylosucrase (NpAS) treatment on sucrose. The greatest CD yield (21.1%, w/w) was achieved from a one-pot dual enzyme reaction at 40 °C for 24 h. The maximum level of CD production (15.1 mg/mL) was achieved with 0.5 M sucrose in a simultaneous mode of dual enzyme reaction, whereas the reaction with 0.1 M sucrose was the most efficient with regard to conversion yield. Consequently, dual enzyme synthesis of CDs was successfully carried out with no need of starch material. This result can be applied as a novel efficient bioconversion process that does not require the high temperature necessary for starch liquefaction by thermostable α-amylase in conventional industrial processing.
- Published
- 2016
- Full Text
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369. An unusual chimeric amylosucrase generated by domain-swapping mutagenesis.
- Author
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Seo DH, Jung JH, Jung DH, Park S, Yoo SH, Kim YR, and Park CS
- Subjects
- Bacterial Proteins chemistry, Deinococcus enzymology, Deinococcus genetics, Enzyme Stability, Glucosyltransferases chemistry, Kinetics, Molecular Dynamics Simulation, Mutagenesis, Neisseria enzymology, Neisseria genetics, Protein Engineering, Protein Structure, Quaternary, Recombinant Fusion Proteins chemistry, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Substrate Specificity, Bacterial Proteins genetics, Bacterial Proteins metabolism, Glucosyltransferases genetics, Glucosyltransferases metabolism
- Abstract
Amylosucrase (ASase; EC 2.4.1.4) synthesizes α-1,4-glucans using sucrose as a sole substrate. The aim of this study was to compare the enzymatic properties of four recombinant ASase genes to determine the underlying mechanisms thereof. Following cloning and expression in Escherichia coli, we determined that the ASase enzyme from Deinococcus geothermalis (DGAS) had the highest thermostability whereas ASase from Neisseria polysaccharea (NPAS) showed the greatest polymerization activity. Chimeric ASases were constructed using dgas and npas genes by overlap extension polymerase chain reaction. Two of the six chimeric ASases generated, NPAS-B' and DGAS-B, showed ASase activity using sucrose as the sole substrate. However, DGAS-B was not able to produce longer α-1,4-glucans; the highest degree of polymerization was <12. In the kinetic study, not only the substrate binding affinity but also the production rate of DGAS-B was greater than those of DGAS. Molecular dynamic computational simulation suggested that DGAS-B could not synthesize longer glucan chains because of the change in flexibilities of loops 4, 7, and 8as compared to those of DGAS., (Copyright © 2016 Elsevier Inc. All rights reserved.)
- Published
- 2016
- Full Text
- View/download PDF
370. Low digestion property of amylosucrase-modified waxy adlay starch.
- Author
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Kim EJ, Kim HR, Choi SJ, Park CS, and Moon TW
- Abstract
Structural and digestion properties of amylosucrase-modified waxy adlay starch were investigated. The unique reaction of amylosucrase caused a decrease and an increase in the proportion of short chains and long chains, respectively, via attachment of glucosyl units to the non-reducing ends of branch chains. The in vitro digestion profile of amylosucrase-modified starch revealed that elongated branch chains were the main reason for high contents of slowly digestible and resistant starches due to formation of a more perfect crystalline structure via easy association between elongated branch chains. The glucose response in mice after consumption of amylosucrase-modified starch was similar to the response for commercial resistant starch with a gradual increase followed by a gradual decrease in blood glucose concentrations over a prolonged time. Both in vitro and in vivo tests were used to verify increased resistance to digestive enzymes caused by amylosucrase modification.
- Published
- 2016
- Full Text
- View/download PDF
371. Enzymatic synthesis of 2-deoxyglucose-containing maltooligosaccharides for tracing the location of glucose absorption from starch digestion.
- Author
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Lee BH, Koh DW, Territo PR, Park CS, Hamaker BR, and Yoo SH
- Subjects
- Animals, Deoxyglucose analysis, Digestion, Glycoside Hydrolase Inhibitors metabolism, Humans, Hydrolysis, Neisseria enzymology, Oligosaccharides analysis, alpha-Amylases metabolism, alpha-Glucosidases metabolism, Deoxyglucose metabolism, Glucose analysis, Glucose metabolism, Ileum physiology, Oligosaccharides metabolism, Starch metabolism
- Abstract
The ileal brake mechanism which induces a potentially beneficial slower gastric emptying rate and increased satiety is triggered by macronutrients including glucose from glycemic carbohydrates. For optimization of this diet-induced health benefit, there is the need for a way to determine the location of glucose deposition in the small intestine. Labeled 2-deoxyglucose (2-DG) can be used to trace the location of glucose absorption due to its accumulative property in the small intestine enterocytes. However, because pure glucose, or 2-DG, is directly absorbed in the proximal small intestine, we designed 2-DG containing maltooligosaccharides (2-DG-MOs) that can be used with a mild α-glucosidase inhibitor to attain an analytical method for determining location-specific delivery of glucose and its physiological effect., (Copyright © 2015 Elsevier Ltd. All rights reserved.)
- Published
- 2015
- Full Text
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372. Flavonoid glucosylation by non-Leloir glycosyltransferases: formation of multiple derivatives of 3,5,7,3',4'-pentahydroxyflavane stereoisomers.
- Author
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Overwin H, Wray V, and Hofer B
- Subjects
- Glycosylation, Stereoisomerism, Flavonoids metabolism, Glycosyltransferases metabolism, Neisseria enzymology, Streptococcus oralis enzymology
- Abstract
Flavonoids are known to possess a multitude of biological activities. Therefore, diversification of the core structures is of considerable interest. One of nature's important tailoring reactions in the generation of bioactive compounds is glycosylation, which is able to influence numerous molecular properties. Here, we examined two non-Leloir glycosyltransferases that use sucrose as an inexpensive carbohydrate donor, glycosyltransferase R from Streptococcus oralis (GtfR) and amylosucrase from Neisseria polysaccharea (Ams), for the glucosylation of flavonoids. Flavones generally were poor substrates. Several inhibited Ams. In contrast, flavanes were well accepted by both enzymes. All glucose attachments occurred via α1 linkages. Comparison of the three available stereoisomers of 3,5,7,3',4'-pentahydroxyflavane revealed significant differences in glycoside formation between them as well as between the two enzymes. The latter were shown to possess largely complementary product ranges. Altogether, three of the four hydroxy substituents of the terminal flavonoid rings were glycosylated. Typically, Ams glucosylated the B ring at position 3', whereas GtfR glucosylated this ring at position 4' and/or the A ring at position 7. In several instances, short carbohydrate chains were attached to the aglycones. These contained α 1-4 linkages when formed by Ams, but α 1-3 bonds when generated by GtfR. The results show that both enzymes are useful catalysts for the glucodiversification of flavanes. In total, more than 16 products were formed, of which seven have previously not been described.
- Published
- 2015
- Full Text
- View/download PDF
373. The structure of amylosucrase in complex with fructose
- Author
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Lars K. Skov, O. Mirza, and Michael Gajhede
- Subjects
Amylosucrase ,chemistry.chemical_compound ,biology ,chemistry ,Biochemistry ,Structural Biology ,biology.protein ,Fructose - Published
- 2002
374. Structural analysis of substrate binding by the glucan synthesizing enzyme amylosucrase
- Author
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O. Mirza, Anette Henriksen, Magali Remaud-Simeon, Patricia Sarçabal, R.-M. Willemot, Michael Gajhede, G. Potocki De Montalk, Lars K. Skov, and Pierre Monsan
- Subjects
chemistry.chemical_classification ,Amylosucrase ,Enzyme ,chemistry ,biology ,Biochemistry ,Structural Biology ,biology.protein ,Substrate (chemistry) ,Protein engineering ,Enzyme catalysis ,Glucan - Published
- 2000
375. Biotransformation of phloretin by amylosucrase yields three novel dihydrochalcone glucosides.
- Author
-
Overwin H, Wray V, and Hofer B
- Subjects
- Biocatalysis drug effects, Biotransformation drug effects, Cell Death drug effects, Cell Line, Tumor, Glycosylation drug effects, Humans, Magnetic Resonance Spectroscopy, Phloretin chemistry, Phloretin toxicity, Time Factors, Chalcones metabolism, Glucosides metabolism, Glucosyltransferases metabolism, Phloretin metabolism
- Abstract
Glycosylation is one of the most important tailoring reactions for natural products. It typically exerts profound direct or indirect effects on their biological activity. The dihydrochalcone phloretin and its known sugar derivatives, particularly phlori(d)zin, have been shown to influence various cellular processes. We found that a non-Leloir glycosyltransferase, amylosucrase from Neisseria polysaccharea, is an excellent catalyst for the stereospecific glucosylation of phloretin at the 4' position. Three novel phloretin derivatives were obtained, the first ones in which the sugar-aglycone bond possesses the configuration. A first biological characterization in a cell viability assay showed that each sugar attachment reduced the compound toxicity approximately two-fold., (Copyright © 2015 Elsevier B.V. All rights reserved.)
- Published
- 2015
- Full Text
- View/download PDF
376. Characterization of enzymatically modified rice and barley starches with amylosucrase at scale-up production.
- Author
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Kim BS, Kim HS, and Yoo SH
- Subjects
- Biocatalysis, Neisseria enzymology, Solubility, Transition Temperature, Viscosity, Amylopectin chemistry, Bacterial Proteins chemistry, Glucosyltransferases chemistry, Hordeum chemistry, Oryza chemistry
- Abstract
Physicochemical properties of Neisseria polysaccharea amylosucrase (NpAS)-treated rice and barley starches were investigated at scale-up production. Pre-gelatinized rice and barley starches were treated with significantly lower NpAS dose (0.1 U/mL) but 100 times larger reaction volume (3500 mL), compared to the analytical scale (35 mL) used in the previous study. NpAS-treated starches in this scale-up production were characterized with respect to reaction efficiency (RE), resistant starch (RS) content, amylopectin (AP) branch-chain length distribution, solubility, swelling power, pasting viscosity, and thermal transition properties. The RE enhanced up to 1.8 times by increasing the reaction volume, which improved the RS content and AP branch-chain lengths of NpAS-treated starches. Compared with the native starch, NpAS-treated starches exhibited lower solubility and swelling power, lower pasting viscosity, and a large increase in the melting peak temperature. Consequently, NpAS treatment of pre-gelatinized starches in this study would be a potential way of replacing commercial RS production., (Copyright © 2015 Elsevier Ltd. All rights reserved.)
- Published
- 2015
- Full Text
- View/download PDF
377. Synthesis and biological evaluation of a novel baicalein glycoside as an anti-inflammatory agent.
- Author
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Kim KH, Park YD, Park H, Moon KO, Ha KT, Baek NI, Park CS, Joo M, and Cha J
- Subjects
- Animals, Antioxidants chemistry, Antioxidants pharmacology, Cell Line, Gene Expression drug effects, Lipopolysaccharides pharmacology, Macrophages drug effects, Macrophages metabolism, Mice, NF-E2-Related Factor 2 metabolism, Nitric Oxide metabolism, Reactive Oxygen Species metabolism, Anti-Inflammatory Agents chemistry, Anti-Inflammatory Agents pharmacology, Flavanones chemistry, Flavanones pharmacology, Glycosides chemistry, Glycosides pharmacology
- Abstract
Baicalein-6-α-glucoside (BG), a glycosylated derivative of baicalein, was synthesized by using sucrose and the amylosucrase of Deinococcus geothermalis and tested for its solubility, chemical stability, and anti-inflammatory activity. BG was 26.3 times more soluble than baicalein and highly stable in buffered solutions and Dulbecco׳s modified Eagle medium containing 10% fetal bovine serum. BG treatment decreased the production of nitric oxide in RAW 264.7 cells treated with lipopolysaccharide (LPS). Luciferase reporter assays, western blots, reverse transcription-polymerase chain reaction, and flow cytometric analyses indicated that BG activated nuclear factor erythroid 2-related factor 2 (Nrf2), an antioxidant transcription factor that confers protection from various inflammatory diseases, induced Nrf2-dependent gene expression, and suppressed the production of reactive oxygen species elicited by LPS more effectively than baicalein. Cellular uptake of BG assessed by confocal microscopy and HPLC analysis of the cell-free extracts of RAW 264.7 cells demonstrated that BG was gradually converted to baicalein inside the cells. These results explain that glycosylation increased the bioavailability of baicalein by helping to protect this vital molecule from chemical or enzymatic oxidation. Therefore, BG, a glycosylated derivative of baicalein, can be an alternative to baicalein as a therapeutic drug., (Copyright © 2014 Elsevier B.V. All rights reserved.)
- Published
- 2014
- Full Text
- View/download PDF
378. Essential role of amino acid position 226 in oligosaccharide elongation by amylosucrase from Neisseria polysaccharea.
- Author
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Cambon E, Barbe S, Pizzut-Serin S, Remaud-Simeon M, and André I
- Subjects
- Amino Acid Substitution, Amino Acids genetics, Catalytic Domain, DNA Mutational Analysis, Glucosyltransferases chemistry, Glucosyltransferases genetics, Kinetics, Models, Molecular, Mutagenesis, Site-Directed, Mutant Proteins chemistry, Mutant Proteins genetics, Mutant Proteins metabolism, Neisseria genetics, Protein Conformation, Sucrose metabolism, Amino Acids metabolism, Glucosyltransferases metabolism, Neisseria enzymology, Oligosaccharides metabolism
- Abstract
Amylosucrase from Neisseria polysaccharea is a remarkable transglucosylase that synthesizes an insoluble amylose-like polymer from sole substrate sucrose. One particular amino acid, Arg226, was proposed from molecular modeling studies to play an important role in the formation of the active site topology and in the accessibility of ligands to the catalytic site. The systematic mutation of this Arg residue by all 19 other possible amino acids revealed that all single-mutants had a higher activity on sucrose compared to the wild-type enzyme. An extensive kinetic investigation showed that catalytic efficiencies are greatly impacted by the presence of natural acceptors in the reaction media, their chain length and the nature of the amino acid at position 226. Compared to the wild-type enzyme, the R226N mutant showed a 10-fold enhancement in the catalytic efficiency and a nearly twofold higher production of an insoluble amylose-like polymer that can be of interest for biotechnological applications., (© 2014 Wiley Periodicals, Inc.)
- Published
- 2014
- Full Text
- View/download PDF
379. Functional characterization of Synechococcus amylosucrase and fructokinase encoding genes discovers two novel actors on the stage of cyanobacterial sucrose metabolism.
- Author
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Perez-Cenci M and Salerno GL
- Subjects
- Fructokinases metabolism, Glucosyltransferases metabolism, Synechococcus enzymology, Synechococcus metabolism, Fructokinases genetics, Gene Expression Regulation, Bacterial, Genes, Bacterial, Glucosyltransferases genetics, Sucrose metabolism, Synechococcus genetics
- Abstract
Plants and most cyanobacteria metabolize sucrose (Suc) with a similar set of enzymes. In Synechococcus sp. PCC 7002, a marine cyanobacterium strain, genes involved in Suc synthesis (spsA and sppA) have been characterized; however, its breakdown was still unknown. Indeed, neither invertase nor sucrose synthase genes, usually found in plants and cyanobacteria, were found in that Synechococcus genome. In the present study, we functionally characterized the amsA gene that codes for an amylosucrase (AMS), a glycoside-hydrolase family 13 enzyme described in bacteria, which may catabolyze Suc in Synechococcus. Additionally, we identified and characterized the frkA gene that codes for a fructokinase (FRK), enzyme that yields fructose-6P, one of the substrates for Suc synthesis. Interestingly, we demonstrate that spsA, sppA, frkA and amsA are grouped in a transcriptional unit that were named Suc cluster, whose expression is increased in response to a salt treatment. This is the first report on the characterization of an AMS and FRK in an oxygenic photosynthetic microorganism, which could be associated with Suc metabolism., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)
- Published
- 2014
- Full Text
- View/download PDF
380. Branch chain elongation by amylosucrase: production of waxy corn starch with a slow digestion property.
- Author
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Kim BK, Kim HI, Moon TW, and Choi SJ
- Subjects
- Biocatalysis, Digestion, Molecular Structure, Bacterial Proteins chemistry, Glucosyltransferases chemistry, Neisseria enzymology, Plant Extracts chemistry, Starch chemistry, Zea mays chemistry
- Abstract
Starches with high slowly digestible starch (SDS) contents were prepared by treating completely gelatinized waxy corn starch with amylosucrase. The structural properties of the prepared starches were then investigated. The content of SDS increased by up to 38.7% after amylosucrase modification, and the portion of chains with degree of polymerisation (DP) 25-36 increased, while the portion of chains with DP⩽12 decreased. Amylosucrase-modified starches showed a weak B-type crystalline structure. A slight increase in the degree of relative crystallinity was observed with increased reaction time. The thermal properties, including melting temperature and enthalpy, of the amylosucrase-modified starches were higher than for the control starch. Although the amylosucrase-modified starches showed varying structural properties according to reaction time (1-45 h), their digestibilities did not change much after 6 h. By controlling the reaction time of the amylosucrase treatment, a tailored starchy food containing the desired amount of SDS can be produced., (Copyright © 2013 Elsevier Ltd. All rights reserved.)
- Published
- 2014
- Full Text
- View/download PDF
381. Probing impact of active site residue mutations on stability and activity of Neisseria polysaccharea amylosucrase.
- Author
-
Daudé D, Topham CM, Remaud-Siméon M, and André I
- Subjects
- Amino Acid Sequence, Amino Acid Substitution, Catalytic Domain genetics, Enzyme Stability, Evolution, Molecular, Genetic Variation, Glucans metabolism, Glucosyltransferases chemistry, Hydrogen Bonding, Models, Molecular, Mutagenesis, Site-Directed, Neisseria genetics, Substrate Specificity, Thermodynamics, Glucosyltransferases genetics, Glucosyltransferases metabolism, Neisseria enzymology
- Abstract
The amylosucrase from Neisseria polysaccharea is a transglucosidase from the GH13 family of glycoside-hydrolases that naturally catalyzes the synthesis of α-glucans from the widely available donor sucrose. Interestingly, natural molecular evolution has modeled a dense hydrogen bond network at subsite -1 responsible for the specific recognition of sucrose and conversely, it has loosened interactions at the subsite +1 creating a highly promiscuous subsite +1. The residues forming these subsites are considered to be likely involved in the activity as well as the overall stability of the enzyme. To assess their role, a structure-based approach was followed to reshape the subsite -1. A strategy based on stability change predictions, using the FoldX algorithm, was considered to identify the best candidates for site-directed mutagenesis and guide the construction of a small targeted library. A miniaturized purification protocol was developed and both mutant stability and substrate promiscuity were explored. A range of 8 °C between extreme melting temperature values was observed and some variants were able to synthesize series of oligosaccharides with distributions differing from that of the parental enzyme. The crucial role of subsite -1 was thus highlighted and the biocatalysts generated can now be considered as starting points for further engineering purposes., (© 2013 The Protein Society.)
- Published
- 2013
- Full Text
- View/download PDF
382. The structure of amylosucrase from Deinococcus radiodurans has an unusual open active-site topology.
- Author
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Skov LK, Pizzut-Serin S, Remaud-Simeon M, Ernst HA, Gajhede M, and Mirza O
- Subjects
- Bacterial Proteins genetics, Bacterial Proteins metabolism, Catalytic Domain, Crystallography, X-Ray, Deinococcus enzymology, Deinococcus genetics, Escherichia coli genetics, Glucans metabolism, Glucosyltransferases genetics, Glucosyltransferases metabolism, Models, Molecular, Protein Binding, Protein Multimerization, Protein Structure, Secondary, Recombinant Proteins chemistry, Recombinant Proteins genetics, Recombinant Proteins metabolism, Substrate Specificity, Sucrose metabolism, Bacterial Proteins chemistry, Deinococcus chemistry, Glucans chemistry, Glucosyltransferases chemistry, Sucrose chemistry
- Abstract
Amylosucrases (ASes) catalyze the formation of an α-1,4-glucosidic linkage by transferring a glucosyl unit from sucrose onto an acceptor α-1,4-glucan. To date, several ligand-bound crystal structures of wild-type and mutant ASes from Neisseria polysaccharea and Deinococcus geothermalis have been solved. These structures all display a very similar overall conformation with a deep pocket leading to the site for transglucosylation, subsite -1. This has led to speculation on how sucrose enters the active site during glucan elongation. In contrast to previous studies, the AS structure from D. radiodurans presented here has a completely empty -1 subsite. This structure is strikingly different from other AS structures, as an active-site-lining loop comprising residues Leu214-Asn225 is found in a previously unobserved conformation. In addition, a large loop harbouring the conserved active-site residues Asp133 and Tyr136 is disordered. The result of the changed loop conformations is that the active-site topology is radically changed, leaving subsite -1 exposed and partially dismantled. This structure provides novel insights into the dynamics of ASes and comprises the first structural support for an elongation mechanism that involves considerable conformational changes to modulate accessibility to the sucrose-binding site and thereby allows successive cycles of glucosyl-moiety transfer to a growing glucan chain.
- Published
- 2013
- Full Text
- View/download PDF
383. Thin-layer chromatography using multiple development for analysis of reaction products of sucrases
- Author
-
Shigeyuki Hamada, Toshio Horikoshi, and Toshihiko Koga
- Subjects
chemistry.chemical_classification ,Sucrose ,Inulosucrase ,Chromatography ,Glycoside Hydrolases ,beta-Fructofuranosidase ,biology ,Chemistry ,Carbohydrates ,Levansucrase ,General Chemistry ,Thin-layer chromatography ,Dextransucrase ,Sucrase ,chemistry.chemical_compound ,Amylosucrase ,Enzyme ,Biochemistry ,Glucosyltransferases ,biology.protein ,Chromatography, Thin Layer - Published
- 1987
384. New Studies on Amylosucrase, a Bacterial α-d-Glucosylase That Directly Converts Sucrose to a Glycogen-like α-Glucan
- Author
-
Edward J. Hehre and Gentaro Okada
- Subjects
chemistry.chemical_classification ,Sucrose ,Glycogen ,biology ,Chemistry ,Fructose ,Cell Biology ,Maltose ,Biochemistry ,carbohydrates (lipids) ,chemistry.chemical_compound ,Amylosucrase ,Amylopectin ,biology.protein ,Maltase ,Molecular Biology ,Glucan - Abstract
Amylosucrase (EC 2.4.1.4) was solubilized and purified more than 100-fold from Neisseria perflava cells. Preparations had a specific activity of about 3 i.u. per mg of protein, were free of maltase and α-glucan phosphorylase activity, and converted sucrose to an amylaceous α-glucan, fructose, and minor amounts of maltosaccharides (Keq = 33, ΔG° = -2.1 Cal). Sugar nucleotide mediation was absent. Added UDP or ADP did not affect the conversion rate, or provide UDP- or ADP-[14C]glucose from [14C]sucrose even when polymerization was prevented by α-amylase. The α-glucan formed from sucrose was glycogen-like (e.g. 48% hydrolyzed to maltose by β-amylase). The branches may be produced in the usual way, assuming the amylosucrase preparations were contaminated with a dextrinyl transferring (branching) enzyme. It is possible, however, that amylosucrase itself may produce the entire glucan structure by a distinctive process involving α-d-glucosyl (monomer) transfer exclusively. Specificity of the enzyme for monomer was apparent as only α-d-glucosyl was transferred from amylopectin, maltoheptaose, and sucrose to [14C]fructose (forming labeled sucrose). The C4-linked α-maltosyl, α-maltotriosyl, etc., terminal segmers of the first two compounds and the α-maltosyl group of 4g α-d-glucosyl-sucrose were not transferred. α-Glucan synthesis by amylosucrase was found to require a primer. Onset of synthesis from sucrose free of amylaceous impurities was long delayed, and this delay was abolished by amylopectin or glycogen, 3 to 6 x 10-6 m end groups. Sucrose, with a donor affinity of Ks = 17 mm, appears to bind nonproductively at the acceptor site since high concentrations greatly depressed α-glucan synthesis (Kss = 0.3 m). In action on [14C]sucrose, amylosucrose produced α-glucan comprising 99.6% [14C]glucose and 0.4% primer; such evidence for [14C]glucose polymerization is lacking in reports of starch synthesis from sucrose by plant enzyme systems.
- Published
- 1974
385. SYNTHESIS OF A POLYSACCHARIDE OF THE STARCH-GLYCOGEN CLASS FROM SUCROSE BY A CELL-FREE, BACTERIAL ENZYME SYSTEM (AMYLOSUCRASE)
- Author
-
Edward J. Hehre
- Subjects
chemistry.chemical_classification ,Sucrose ,biology ,Glycogen ,Starch ,Cell Biology ,Cell free ,Polysaccharide ,biology.organism_classification ,Biochemistry ,chemistry.chemical_compound ,Amylosucrase ,chemistry ,Enzyme system ,biology.protein ,Molecular Biology ,Bacteria - Published
- 1949
386. An enzymic synthesis of a sucrose analog: α-d-xylopyranosyl-β-d-fructofuranoside
- Author
-
David Sidney Feingold, Shlomo Hestrin, and Gad Avigad
- Subjects
Sucrose ,biology ,Disaccharide ,Levansucrase ,General Medicine ,Xylose ,Disaccharides ,Dextransucrase ,Sucrase ,chemistry.chemical_compound ,Amylosucrase ,Invertase ,chemistry ,Biochemistry ,biology.protein ,Organic Chemicals ,Raffinose - Abstract
Levansucrase transferred the β-fructofuranosyl group of raffinose to the anomeric carbon of xylose. The disaccharide “xylsucrose” formed by this reaction has been isolated and is shown to be α- d -xylopyranosyl-β- d -fructofuranoside. Xylsucrose was hydrolyzed by ordinary yeast invertase and was utilized both as donor and acceptor by levansucrase. Some other sucrases (dextransucrase, amylosucrase, and a special yeast sucrase) were inert towards this sucrose analog. The substrate specificity of levansucrase and the possible role of sucrose analogs in biological polymer synthesis are discussed.
- Published
- 1956
387. Glycogen synthesis by amylosucrase from Neisseria perflava
- Author
-
K. G. Johnson, I. J. McDonald, and C. R. MacKenzie
- Subjects
Sucrose ,Immunology ,Fructose ,Polysaccharide ,Applied Microbiology and Biotechnology ,Microbiology ,chemistry.chemical_compound ,Amylosucrase ,Drug Stability ,Genetics ,Moiety ,Glycogen synthase ,Molecular Biology ,chemistry.chemical_classification ,Neisseria perflava ,Cell-Free System ,biology ,Articles ,General Medicine ,Hydrogen-Ion Concentration ,Enzyme ,chemistry ,Biochemistry ,Glucosyltransferases ,biology.protein ,Neisseria ,Glycogen - Abstract
Amylosucrase (sucrose: 1,4-α-D-glucan 4-α-glucosyltransferase; EC 2.4.1.4) which mediates the transfer of the glucosyl moiety of sucrose to a growing α-1,4-glucan chain is a constitutive enzyme of Neisseria perflava. The products of enzymic action are insoluble glycogenlike polysaccharides and fructose, the latter being a competitive inhibitor of the enzyme (Ki = 20 mM). The enzyme is extremely stable and appears to bind very tightly to its polymerized product. Properties of product-bound enzyme reflect those of the native complex.
- Published
- 1977
388. Neisseria perflava amylosucrase: characterization of its product polysaccharide and a study of its inhibition by sucrose derivatives
- Author
-
Peter J. Reilly, Bernard Y. Tao, and John F. Robyt
- Subjects
chemistry.chemical_classification ,Sucrose ,biology ,Molecular mass ,Glycogen ,Chemistry ,Organic Chemistry ,Polysaccharides, Bacterial ,Substrate (chemistry) ,General Medicine ,Polysaccharide ,Biochemistry ,Analytical Chemistry ,Amylosucrase ,chemistry.chemical_compound ,Kinetics ,Structure-Activity Relationship ,Glucosyltransferases ,biology.protein ,Maltotriose ,Glycosyl ,Neisseria - Abstract
Neisseria perflava amylosucrase forms from sucrose a polysaccharide very similar to glycogen, except that a larger proportion of its d -glucosyl residues are in short branches. Iodine staining of samples taken during polysaccharide formation indicate that the initial product is less branched than that formed at longer times. This glycogen-like polysaccharide has an estimated molecular mass range of 1 MD to 20 MD. Sucrose derivatives modified at C-3 (3-deoxysucrose and α- d -allopyranosyl β- d -fructofuranoside), C-6 (6-deoxysucrose and 6-deoxy-6-fluorosucrose), and both C-4 and C-6 (4,6-dideoxysucrose) were tested as inhibitors of amylosucrase. Derivatives modified at C-6 were potent competitive inhibitors, with K i values of 6.2 ±0.3m m (6-deoxysucrose) and 0.50 ±0.06m m (6-deoxy-6-fluorosucrose). The K M value of sucrose is 26.5 ±4.6m m . Sucrose derivatives modified at C-3 were not significantly inhibitory over the concentration range tested. 4,6-Dideoxysucrose gave an unusual, non-competitive inhibition, in that, increasing its concentration did not produce a commensurate increase in the level of inhibition, which instead appeared to approach a limit. None of these sucrose derivatives was a substrate for amylosucrase, nor were they glycosyl donors to maltotriose.
- Published
- 1988
389. Glycogen metabolism in the genus Neisseria : synthesis from sucrose by amylosucrase
- Author
-
C. R. MacKenzie, I. J. McDonald, and K. G. Johnson
- Subjects
Sucrose ,Immunology ,Fructose ,Biology ,Polysaccharide ,Applied Microbiology and Biotechnology ,Microbiology ,Binding, Competitive ,chemistry.chemical_compound ,Amylosucrase ,Genetics ,Glycogen branching enzyme ,Molecular Biology ,Polyacrylamide gel electrophoresis ,chemistry.chemical_classification ,Glycogen ,General Medicine ,biology.organism_classification ,Molecular biology ,Kinetics ,chemistry ,Biochemistry ,Glucosyltransferases ,biology.protein ,Electrophoresis, Polyacrylamide Gel ,Neisseria - Abstract
Eight strains (seven species) of Neisseria were found to possess intracellular amylosucrases capable of synthesizing glycogen directly from sucrose. All eight systems were stimulated by primer glycogen, possessed similar kinetic parameters, and were competitively inhibited to similar degrees by fructose. The enzymes bound tightly to their polysaccharide products but these complexes could be readily dissociated by polyacrylamide gel electrophoresis. Some of the enzyme–product complexes appeared to be virtually free of contaminating proteins.
- Published
- 1978
390. De novo synthesis of glycosidic linkages by glycosylases: utilization of -D-glucopyranosyl fluoride by amylosucrase
- Author
-
Gentaro Okada and Edward J. Hehre
- Subjects
chemistry.chemical_classification ,biology ,Organic Chemistry ,Glycosidic bond ,General Medicine ,Fructose ,Biochemistry ,Analytical Chemistry ,De novo synthesis ,chemistry.chemical_compound ,Amylosucrase ,Fluorides ,Glucose ,chemistry ,DNA glycosylase ,Glucosyltransferases ,Polysaccharides ,biology.protein ,Organic chemistry ,Fluoride - Published
- 1973
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