DOCSLIB.ORG

Comparative Study on Amylosucrases Derived from Deinococcus Species

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Comparative Study on Amylosucrases Derived from Deinococcus Species Amylase 2019; 3: 19–31 Research Article Ki-Tae Kim, Chan-Su Rha, Young Sung Jung, Ye-Jin Kim, Dong-Hyun Jung, Dong-Ho Seo, Cheon-Seok Park* Comparative study on amylosucrases derived from Deinococcus species and catalytic characterization and use of amylosucrase derived from Deinococcus wulumuqiensis https://doi.org/10.1515/amylase-2019-0002 received December 24, 2018; accepted May 16, 2019. 8 in ASases may affect the acceptor specificity of ASases and result in a distinctive acceptor specificity of DWAS. Abstract: Amylosucrase (ASase; EC 2.4.1.4), a versatile enzyme, exhibits three characteristic activities: Keywords: amylosucrase; Deinococcus; isovitexin; hydrolysis, isomerization, and transglycosylation. In transglycosylation. this study, a novel ASase derived from Deinococcus wulumuquiensis (DWAS) was identified and expressed in Escherichia coli. The optimal reaction temperature and pH for the sucrose hydrolysis activity of DWAS were Abbreviations determined to be 45 °C and 9.0, respectively. DWAS ASase, amylosucrase; CD, cyclodextrin; BCA, bicinchoninic displays relatively high thermostability compared with acid; CR, conserved region; DNS, dinitrosalicylic other ASases, as demonstrated by half-life of 96.7 and acid; DSF, differential scanning fluorimetry; DGAS, 4.7 min at 50 °C and 55 °C, respectively. DWAS fused with amylosucrase from Deinococcus geotermalis; DRAS, 6×His was successfully purified to apparent homogeneity amylosucrase from Deinococcus radiodurans; DRpAS, with a molecular mass of approximately 72 kDa by amylosucrase from Deinococcus radiopugnans; DWAS, Ni-NTA affinity chromatography and confirmed by SDS- amylosucrase from Deinococcus wulumuqiensis; GH, PAGE. DWAS transglycosylation activity can be used to glycoside hydrolase; HPAEC, high-performance anion- modify isovitexin, a representative flavone C-glucoside exchange chromatography; HPLC, high-performance contained in buckwheat sprouts to increase its limited liquid chromatography; LC/MS liquid chromatography/ bioavailability, which is due to its low absorption rate and mass spectrometry; MD, molecular dynamics; NPAS, unstable structure in the human body. Using isovitexin amylosucrase from Neisseria polysaccharea; RT, retention as a substrate, the major transglycosylation product of time. DWAS was found to be isovitexin monoglucoside. The comparison of transglycosylation reaction products of DWAS with those of other ASases derived from Deinococcus species revealed that the low sequence homology of loop 1 Introduction Amylosucrase (ASase; EC 2.4.1.4) is a hydrolytic enzyme belonging to glycoside hydrolase (GH) family GH13. *Corresponding author: Cheon-Seok Park, Graduate School of Evolutionary relationships between 152 GH13 enzymes Biotechnology and Institute of Life Science and Resources, Kyung Hee revealed that ASase together with sucrose hydrolases University, Yongin 17140, Republic of Korea, E-mail: [email protected] Ki-Tae Kim, Chan-Su Rha, Young Sung Jung, Ye-Jin Kim, and sucrose phosphorylases occupied a position Dong-Hyun Jung, Graduate School of Biotechnology and Institute in the tree between the ‘oligo-1,6-glucosidases’ and of Life Science and Resources, Kyung Hee University, Yongin 17140, ‘neopullulanases’ [1]. Previous in silico analysis of GH13 Republic of Korea enzymes assigned ASase to subfamily GH13_4 with Dong-Ho Seo, Research Group of Healthcare, Korea Food Research having both sucrose hydrolysis activity and the “QpDln” Institute, Wanju 55365, Republic of Korea Open Access. © 2019 Ki-Tae Kim et al. et al., published by De Gruyter. This work is licensed under the Creative Commons Attribution alone 4.0 License. 20 Ki-Tae Kim et al. sequence as a characteristic of the subfamily of oligo- the major flavone C-glycosides present in mung bean and 1,6-glucosidase [1,2]. While its sucrose hydrolysis activity buckwheat sprouts [28-30]. It acts as a genetic mediator in involves cleaving the α-1,β-2-glycosidic linkage in sucrose, innate and acquired immune mechanisms. Isovitexin has the enzyme exhibits two additional characteristic catalytic been found to exert anti-inflammatory and antioxidant activities, i.e., isomerase activity that produces sucrose activities by inhibiting the MAPK and NFκB pathways [31-33]. isomers (turanose and trehalulose) and transglycosylation In addition to these activities, various health benefits, such activity that produces α-1,4-glucans, which are amylose- as anti-cancer, anti-estrogenic, and anti-diabetes benefits like polymers [3-5]. In this so-called “transglycosylation” have been reported [34-37]. However, isovitexin displays reaction, the enzyme transfers the glucosyl moiety from both low solubility and stability [38]. During the digestion sucrose to glucose or fructose resulting from sucrose process of flavonoids in the body, C-multiglucosides are hydrolysis. Recently, ASase transglycosylation activity constantly absorbed in the intestine and distributed has been extensively studied for various biotechnological to other tissues, whereas C-monoglucosides are not applications because the enzyme can transfer glucose absorbed well and are unstable due to deglycosylation from an inexpensive sucrose substrate without requiring or degradation by the human intestinal bacteria present the relatively expensive UDP-glucose as a substrate [6]. in the large intestine [33,39]. Therefore, the application ASase was first studied in the Neisseria genus, and of isovitexin in its native C-monoglucoside state remains studies on ASase have involved the synthesis of glycogen- limited in the pharmaceutical and food industries. Several like α-glucan from sucrose [6]. Tao et al. [7] reported the strategies including the use of cyclodextrin (CD) [40], characterization of glycogen-like polysaccharides generated chemical synthesis of glycosides [41], and enzymatic by Neisseria perflava ASase and ASase inhibition by sucrose transglycosylation [42], could be implemented to overcome derivatives. After performing cloning and characterization this limitation. However, among these strategies, the use of the gene corresponding to ASase derived from Neisseria of CD is not favoured due to its poor water solubility, polysaccharea (NPAS), research mainly focused on its potential formation of crystalline complexes, and catalytic properties, three-dimensional structure, and nephrotoxicity associated with parenteral administration product analysis [8-10] until the investigation of ASases [43]. Although chemical synthesis of glycosides is widely in the Deinococcus genus [11]. Recently, ASases have been used, its drawback entails the protection and selective identified in many microorganisms including Alteromonas deprotection steps required to avoid the formation of side macleodii, Arthrobacter chlorophenolicus, Cellulomonas reactions [44-46]. On the other hand, enzyme glycosylation carbonis, Methylobacillus flagellates, Methylomicrobium presents important advantages. It is an environmentally alcaliphilum, and Synechococcus sp. [12-21]. Among these, friendly method that typically does not require heavy Deinococcus species have been thoroughly studied in the metals and toxic components in the reaction. Furthermore, fields of biotechnology and bioremediation because of the biocatalytic reaction facilitates complete control over their exceptional ability to grow and metabolize under anomeric configuration and high regioselectivity without harsh conditions [22]. Recently, we identified ASases in any protecting groups [47,48]. Therefore, the enzymatic various species of the Deinococcus genus and found that glycosylation strategy has been intensively employed for most species contain ASases in their genomes (data not the synthesis of glycosylated natural products. shown). In this study, we aimed to investigate and analyze Flavonoids are divided into several classes, i.e., the distribution of ASases in 30 Deinococcus species. flavones, flavonols, flavonones, isoflavones, flavan-3-ols, Among the analyzed ASases, a putative ASase gene and anthocyanins, according to their backbone structure derived from Deinococcus wulumuqiensis (DWAS) was correlated to the locations of their B-ring and C-ring cloned and expressed in Escherichia coli. The enzymatic and the chemical structure based on the saturation, properties of recombinant DWAS were examined, and its oxidation, and hydroxylation of C-rings [23-25]. Almost transglycosylation activity was used to modify isovitexin. all natural flavonoids exist in the form of O-glycosides The product formation of DWAS was compared with or C-glycosides rather than aglycones [25], with the most those of other ASases derived from other Deinococcus common flavonoid structures in plants being flavone species including ASases of Deinococcus geotermalis glycosides and flavonol glycoside [26]. In nature, C-linked (DGAS) and Deinococcus radiopugnans (DRpAS). The glycosylation is mostly found in the flavone group. expansive reactant and product profiles of Deinococcus Among various flavone C-glycosides, isovitexin ASases suggest that the enzyme has a broad acceptor (apigenin 6-C-glucoside) has been more actively studied specificity and potential applications for selective use in due to its biological functionality [27]. Isovitexin is one of transglycosylation. Amylosucrase from Deinococcus wulumuqiensis 21 2 Materials and methods When the optical density at 600 nm reached 0.5-0.6 as measured using a spectrophotometer (Beckman DU 730; Beckman Coulter, Fullerton, CA, USA), 1 mM isopropyl 2.1 Bacterial strains and cloning vectors β-d-1-thiogalactopyranoside
Load more

Recommended publications
  • Bacteria Belonging to Pseudomonas Typographi Sp. Nov. from the Bark Beetle Ips Typographus Have Genomic Potential to Aid in the Host Ecology
    insects Article Bacteria Belonging to Pseudomonas typographi sp. nov. from the Bark Beetle Ips typographus Have Genomic Potential to Aid in the Host Ecology Ezequiel Peral-Aranega 1,2 , Zaki Saati-Santamaría 1,2 , Miroslav Kolaˇrik 3,4, Raúl Rivas 1,2,5 and Paula García-Fraile 1,2,4,5,* 1 Microbiology and Genetics Department, University of Salamanca, 37007 Salamanca, Spain; [email protected] (E.P.-A.); [email protected] (Z.S.-S.); [email protected] (R.R.) 2 Spanish-Portuguese Institute for Agricultural Research (CIALE), 37185 Salamanca, Spain 3 Department of Botany, Faculty of Science, Charles University, Benátská 2, 128 01 Prague, Czech Republic; [email protected] 4 Laboratory of Fungal Genetics and Metabolism, Institute of Microbiology of the Academy of Sciences of the Czech Republic, 142 20 Prague, Czech Republic 5 Associated Research Unit of Plant-Microorganism Interaction, University of Salamanca-IRNASA-CSIC, 37008 Salamanca, Spain * Correspondence: [email protected] Received: 4 July 2020; Accepted: 1 September 2020; Published: 3 September 2020 Simple Summary: European Bark Beetle (Ips typographus) is a pest that affects dead and weakened spruce trees. Under certain environmental conditions, it has massive outbreaks, resulting in attacks of healthy trees, becoming a forest pest. It has been proposed that the bark beetle’s microbiome plays a key role in the insect’s ecology, providing nutrients, inhibiting pathogens, and degrading tree defense compounds, among other probable traits. During a study of bacterial associates from I. typographus, we isolated three strains identified as Pseudomonas from different beetle life stages. In this work, we aimed to reveal the taxonomic status of these bacterial strains and to sequence and annotate their genomes to mine possible traits related to a role within the bark beetle holobiont.
    [Show full text]
  • Stereoselective Synthesis of a 4- -Glucoside of Valienamine and Its X
    www.nature.com/scientificreports OPEN Stereoselective synthesis of a 4‑⍺‑glucoside of valienamine and its X‑ray structure in complex with Streptomyces coelicolor GlgE1‑V279S Anshupriya Si1,4, Thilina D. Jayasinghe2,4, Radhika Thanvi1, Sunayana Kapil3, Donald R. Ronning2* & Steven J. Sucheck1* Glycoside hydrolases (GH) are a large family of hydrolytic enzymes found in all domains of life. As such, they control a plethora of normal and pathogenic biological functions. Thus, understanding selective inhibition of GH enzymes at the atomic level can lead to the identifcation of new classes of therapeutics. In these studies, we identifed a 4‑⍺‑glucoside of valienamine (8) as an inhibitor of Streptomyces coelicolor (Sco) GlgE1‑V279S which belongs to the GH13 Carbohydrate Active EnZyme family. The results obtained from the dose–response experiments show that 8 at a concentration of 1000 µM reduced the enzyme activity of Sco GlgE1‑V279S by 65%. The synthetic route to 8 and a closely related 4‑⍺‑glucoside of validamine (7) was achieved starting from readily available D‑maltose. A key step in the synthesis was a chelation‑controlled addition of vinylmagnesium bromide to a maltose‑derived enone intermediate. X‑ray structures of both 7 and 8 in complex with Sco GlgE1‑ V279S were solved to resolutions of 1.75 and 1.83 Å, respectively. Structural analysis revealed the valienamine derivative 8 binds the enzyme in an E2 conformation for the cyclohexene fragment. Also, the cyclohexene fragment shows a new hydrogen‑bonding contact from the pseudo‑diaxial C(3)–OH to the catalytic nucleophile Asp 394 at the enzyme active site.
    [Show full text]
  • Towards the Molecular Understanding of Glycogen Elongation by Amylosucrase
    Towards the molecular understanding of glycogen elongation by amylosucrase. Cécile Albenne, Lars K Skov, Vinh Tran, Michael Gajhede, Pierre Monsan, Magali Remaud Simeon, Gwenaëlle André-Leroux To cite this version: Cécile Albenne, Lars K Skov, Vinh Tran, Michael Gajhede, Pierre Monsan, et al.. Towards the molecular understanding of glycogen elongation by amylosucrase.. Proteins - Structure, Function and Bioinformatics, Wiley, 2007, 66 (1), pp.118-26. 10.1002/prot.21083. hal-02663870 HAL Id: hal-02663870 https://hal.inrae.fr/hal-02663870 Submitted on 31 May 2020 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. Distributed under a Creative Commons Attribution - ShareAlike| 4.0 International License PROTEINS: Structure, Function, and Bioinformatics 66:118–126 (2007) Towards the Molecular Understanding of Glycogen Elongation by Amylosucrase Ce´cile Albenne,1 Lars K. Skov,2 Vinh Tran,3 Michael Gajhede,2 Pierre Monsan,4 Magali Remaud-Sime´on,4* and Gwe´nae¨lle Andre´-Leroux5 1Laboratoire Surfaces Cellulaires et Signalisation chez les Ve´ge´taux, UMR 5546
    [Show full text]
  • Flavonoid Glucodiversification with Engineered Sucrose-Active Enzymes Yannick Malbert
    Flavonoid glucodiversification with engineered sucrose-active enzymes Yannick Malbert To cite this version: Yannick Malbert. Flavonoid glucodiversification with engineered sucrose-active enzymes. Biotechnol- ogy. INSA de Toulouse, 2014. English. NNT : 2014ISAT0038. tel-01219406 HAL Id: tel-01219406 https://tel.archives-ouvertes.fr/tel-01219406 Submitted on 22 Oct 2015 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. Last name: MALBERT First name: Yannick Title: Flavonoid glucodiversification with engineered sucrose-active enzymes Speciality: Ecological, Veterinary, Agronomic Sciences and Bioengineering, Field: Enzymatic and microbial engineering. Year: 2014 Number of pages: 257 Flavonoid glycosides are natural plant secondary metabolites exhibiting many physicochemical and biological properties. Glycosylation usually improves flavonoid solubility but access to flavonoid glycosides is limited by their low production levels in plants. In this thesis work, the focus was placed on the development of new glucodiversification routes of natural flavonoids by taking advantage of protein engineering. Two biochemically and structurally characterized recombinant transglucosylases, the amylosucrase from Neisseria polysaccharea and the α-(1→2) branching sucrase, a truncated form of the dextransucrase from L. Mesenteroides NRRL B-1299, were selected to attempt glucosylation of different flavonoids, synthesize new α-glucoside derivatives with original patterns of glucosylation and hopefully improved their water-solubility.
    [Show full text]
  • New Computational Approaches for Investigating the Impact of Mutations on the Transglucosylation Activity of Sucrose Phosphorylase Enzyme
    Thesis submitted to the Université de la Réunion for award of Doctor of Philosophy in Sciences speciality in bioinformatics New computational approaches for investigating the impact of mutations on the transglucosylation activity of sucrose phosphorylase enzyme Mahesh VELUSAMY Thesis to be presented on 18th December 2018 in front of the jury composed of Mme Isabelle ANDRE, Directeur de Recherche CNRS, INSA de TOULOUSE, Rapporteur M. Manuel DAUCHEZ, Professeur, Université de Reims Champagne Ardennes, Rapporteur M. Richard DANIELLOU, Professeur, Université d'Orléans, Examinateur M. Yves-Henri SANEJOUAND, Directeur de Recherche CNRS, Université de Nantes, Examinateur Mme Irène MAFFUCCI, Maître de Conférences, Université de Technologie de Compiègne, Examinateur Mme Corinne MIRAL, Maître de Conférences HDR, Université de Nantes, Examinateur M. Frédéric CADET, Professeur, Université de La Réunion, Co-directeur de thèse M. Bernard OFFMANN, Professeur, Université de Nantes, Directeur de thèse ெ்்ந்ி ிைு்ூுத் ெ்யாம் ெ்த உதி்ு ையகு் ானகு் ஆ்ற் அிு. -ிு்ு் ுதி் அ்ப் ுுக், எனு த்ைத ேுாி, தா் க்ூி, அ்ண் ு்ு்ுமா், த்ைக பா்பா, ஆ்தா ுு்மா், ீனா, அ்ண் ்ுக், ெபிய்பா, ெபிய்மா ம்ு் உுுைணயா் இு்த அைண்ு ந்ப்கு்ு் எனு மனமா்்த ந்ி. இ்த ஆ்ி்ைக ுுைம அை3த்ு ுுுத்்காரண், எனு ஆ்ி்ைக இய்ுன் ேபராிிய் ெப்னா்் ஆஃ்ேம். ப்ேு துண்கி் நா் மனதாு், ெபாுளாதார அளிு் க்3்ி் இு்தேபாு, என்ு இ்ெனாு த்ைதயாகே இு்ு எ்ைன பா்்ு்ெகா்3ா். ுி்பாக, எனு ூ்றா் ஆ்ு இுிி், அ்்ு எ்ளோ தி்ப்3 க3ைமக் ம்ு் ிர்ிைனக் இு்தாு், அைத்ெபாு்பு்தாு, அ் என்ு ெ்த ெபாுளாதார உதி, ப்கைB்கழக பிு ம்ு் இதர ி்ாக ்ப்த்ப்3 உதிகு்ு எ்ன ைகமா்ு ெகாு்தாு் ஈ3ாகாு.
    [Show full text]
  • Supporting Information High-Throughput Virtual Screening
    Supporting Information High-Throughput Virtual Screening of Proteins using GRID Molecular Interaction Fields Simone Sciabola, Robert V. Stanton, James E. Mills, Maria M. Flocco, Massimo Baroni, Gabriele Cruciani, Francesca Perruccio and Jonathan S. Mason Contents Table S1 S2-S21 Figure S1 S22 * To whom correspondence should be addressed: Simone Sciabola, Pfizer Research Technology Center, Cambridge, 02139 MA, USA Phone: +1-617-551-3327; Fax: +1-617-551-3117; E-mail: [email protected] S1 Table S1. Description of the 990 proteins used as decoy for the Protein Virtual Screening analysis. PDB ID Protein family Molecule Res. (Å) 1n24 ISOMERASE (+)-BORNYL DIPHOSPHATE SYNTHASE 2.3 1g4h HYDROLASE 1,3,4,6-TETRACHLORO-1,4-CYCLOHEXADIENE HYDROLASE 1.8 1cel HYDROLASE(O-GLYCOSYL) 1,4-BETA-D-GLUCAN CELLOBIOHYDROLASE I 1.8 1vyf TRANSPORT PROTEIN 14 KDA FATTY ACID BINDING PROTEIN 1.85 1o9f PROTEIN-BINDING 14-3-3-LIKE PROTEIN C 2.7 1t1s OXIDOREDUCTASE 1-DEOXY-D-XYLULOSE 5-PHOSPHATE REDUCTOISOMERASE 2.4 1t1r OXIDOREDUCTASE 1-DEOXY-D-XYLULOSE 5-PHOSPHATE REDUCTOISOMERASE 2.3 1q0q OXIDOREDUCTASE 1-DEOXY-D-XYLULOSE 5-PHOSPHATE REDUCTOISOMERASE 1.9 1jcy LYASE 2-DEHYDRO-3-DEOXYPHOSPHOOCTONATE ALDOLASE 1.9 1fww LYASE 2-DEHYDRO-3-DEOXYPHOSPHOOCTONATE ALDOLASE 1.85 1uk7 HYDROLASE 2-HYDROXY-6-OXO-7-METHYLOCTA-2,4-DIENOATE 1.7 1v11 OXIDOREDUCTASE 2-OXOISOVALERATE DEHYDROGENASE ALPHA SUBUNIT 1.95 1x7w OXIDOREDUCTASE 2-OXOISOVALERATE DEHYDROGENASE ALPHA SUBUNIT 1.73 1d0l TRANSFERASE 35KD SOLUBLE LYTIC TRANSGLYCOSYLASE 1.97 2bt4 LYASE 3-DEHYDROQUINATE DEHYDRATASE
    [Show full text]
  • The Influence of Starch Modification with Amylosucrase Treatment On
    processes Article The Influence of Starch Modification with Amylosucrase Treatment on Morphological Features Hyeyoung Lee 1 and Inmyoung Park 2,* 1 Division of Applied Bioengineering, Dong-eui University, Busan 47340, Korea; [email protected] 2 Division of Food and Culinary Arts, Youngsan University, Busan 48015, Korea * Correspondence: [email protected]; Tel.: +82-51-540-7236 Received: 22 October 2020; Accepted: 3 November 2020; Published: 4 November 2020 Abstract: Amylosucrase (AS) is a starch-modifying enzyme from Neisseria polysaccharea used to produce low-glycemic starches such as slowly digestible starch (SDS) and resistant starch (RS). The morphology of native, control, and AS-modified waxy corn starches (230 and 460 U) was examined using a particle size analyzer and field-emission scanning electron microscopy (FE-SEM). AS modification of the starch elongated the glucose and resulted in higher SDS and RS contents. The mean particle sizes of the control, 230 U-AS-, and 460 U-AS-treated starches were 56.6 µm, 128.0 µm, and 176.5 µm, respectively. The surface of the 460 U-AS-treated starch was entirely porous and coral-like, while the 230 U-AS-treated starch had a partial dense and flat surface which did not react with AS. FE-SEM of the granule cross section confirmed that the center contained a dense and flat region without any evidence of AS reaction to either of the AS-treated starches. It was assumed that the particle size and porous and sponge-like particle features might be related to the SDS and RS fractions. Keywords: amylosucrase-modified starch; particle size distribution; cross-sectioned particle; morphology; low glycemic starch 1.
    [Show full text]
  • アミロース製造に利用する酵素の開発と改良 Phosphorylase and Muscle Phosphorylase B
    125 J. Appl. Glycosci., 54, 125―131 (2007) !C 2007 The Japanese Society of Applied Glycoscience Proceedings of the Symposium on Amylases and Related Enzymes, 2006 Developing and Engineering Enzymes for Manufacturing Amylose (Received December 5, 2006; Accepted January 5, 2007) Michiyo Yanase,1,＊ Takeshi Takaha1 and Takashi Kuriki1 1 Biochemical Research Laboratory, Ezaki Glico Co., Ltd. (4 ―6 ―5, Utajima, Nishiyodogawa-ku, Osaka 555-8502, Japan) Abstract: Amylose is a functional biomaterial and is expected to be used for various industries. However at present, manufacturing of amylose is not done, since the purification of amylose from starch is very difficult. It has been known that amylose can be produced in vitro by using α-glucan phosphorylases. In order to ob- tain α-glucan phosphorylase suitable for manufacturing amylose, we isolated an α-glucan phosphorylase gene from Thermus aquaticus and expressed it in Escherichia coli. We also obtained thermostable α-glucan phos- phorylase by introducing amino acid replacement onto potato enzyme. α-Glucan phosphorylase is suitable for the synthesis of amylose; the only problem is that it requires an expensive substrate, glucose 1-phosphate. We have avoided this problem by using α-glucan phosphorylase either with sucrose phosphorylase or cellobiose phosphorylase, where inexpensive raw material, sucrose or cellobiose, can be used instead. In these combined enzymatic systems, α-glucan phosphorylase is a key enzyme. This paper summarizes our work on engineering practical α-glucan phosphorylase for industrial processes and its use in the enzymatic synthesis of essentially linear amylose and other glucose polymers. Key words: amylose, glucose polymer, α-glucan phosphorylase, glycogen debranching enzyme, protein engi- neering α-1,4 glucan is the major form of energy reserve from Enzymes for amylose synthesis.
    [Show full text]
  • Ep 1 117 822 B1
    Europäisches Patentamt (19) European Patent Office & Office européen des brevets (11) EP 1 117 822 B1 (12) EUROPÄISCHE PATENTSCHRIFT (45) Veröffentlichungstag und Bekanntmachung des (51) Int Cl.: Hinweises auf die Patenterteilung: C12P 19/18 (2006.01) C12N 9/10 (2006.01) 03.05.2006 Patentblatt 2006/18 C12N 15/54 (2006.01) C08B 30/00 (2006.01) A61K 47/36 (2006.01) (21) Anmeldenummer: 99950660.3 (86) Internationale Anmeldenummer: (22) Anmeldetag: 07.10.1999 PCT/EP1999/007518 (87) Internationale Veröffentlichungsnummer: WO 2000/022155 (20.04.2000 Gazette 2000/16) (54) HERSTELLUNG VON POLYGLUCANEN DURCH AMYLOSUCRASE IN GEGENWART EINER TRANSFERASE PREPARATION OF POLYGLUCANS BY AMYLOSUCRASE IN THE PRESENCE OF A TRANSFERASE PREPARATION DES POLYGLUCANES PAR AMYLOSUCRASE EN PRESENCE D’UNE TRANSFERASE (84) Benannte Vertragsstaaten: (56) Entgegenhaltungen: AT BE CH CY DE DK ES FI FR GB GR IE IT LI LU WO-A-00/14249 WO-A-00/22140 MC NL PT SE WO-A-95/31553 (30) Priorität: 09.10.1998 DE 19846492 • OKADA, GENTARO ET AL: "New studies on amylosucrase, a bacterial.alpha.-D-glucosylase (43) Veröffentlichungstag der Anmeldung: that directly converts sucrose to a glycogen- 25.07.2001 Patentblatt 2001/30 like.alpha.-glucan" J. BIOL. CHEM. (1974), 249(1), 126-35, XP000867741 (73) Patentinhaber: Südzucker AG Mannheim/ • BUTTCHER, VOLKER ET AL: "Cloning and Ochsenfurt characterization of the gene for amylosucrase 68165 Mannheim (DE) from Neisseria polysaccharea: production of a linear.alpha.-1,4-glucan" J. BACTERIOL. (1997), (72) Erfinder: 179(10), 3324-3330, XP002129879 • GALLERT, Karl-Christian • DE MONTALK, G. POTOCKI ET AL: "Sequence D-61184 Karben (DE) analysis of the gene encoding amylosucrase • BENGS, Holger from Neisseria polysaccharea and D-60598 Frankfurt am Main (DE) characterization of the recombinant enzyme" J.
    [Show full text]
  • Enzymatic Production of Human Milk Oligosaccharides
    Downloaded from orbit.dtu.dk on: Dec 20, 2017 Enzymatic production of human milk oligosaccharides Guo, Yao; Mikkelsen, Jørn Dalgaard; Jers, Carsten Publication date: 2014 Document Version Publisher's PDF, also known as Version of record Link back to DTU Orbit Citation (APA): Guo, Y., Mikkelsen, J. D., & Jers, C. (2014). Enzymatic production of human milk oligosaccharides. Technical University of Denmark, Department of Chemical Engineering. General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights. • Users may download and print one copy of any publication from the public portal for the purpose of private study or research. • You may not further distribute the material or use it for any profit-making activity or commercial gain • You may freely distribute the URL identifying the publication in the public portal If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim. Enzymatic production of human milk oligosaccharides Yao Guo Ph.D. Thesis April 2014 Enzymatic production of human milk oligosaccharides Yao Guo Ph.D. thesis, April 2014 Center for BioProcess Engineering 1 Copyright©: Yao Guo April 2014 Center For Bioprocess Engineering Department of Chemical and Biochemical Engineering Technical University of Denmark Søltofts Plads, Building 229 DK-2800 Kgs. Lyngby Denmark Phone: +45 4525 2800 Fax: +45 4525 2258 Web: www.bioeng.kt.dtu.dk Print: J&R Frydenberg A/S København September 2014 ISBN: 978-87-93054-43-1 2 Preface This thesis comprises the research carried out during my PhD study at the Center for BioProcess Engineering, Department of Chemical and Biochemical Engineering, Technical University of Denmark, from August 2010 to April 2014.
    [Show full text]
  • (12) Patent Application Publication (10) Pub. No.: US 2011/0165635 A1 Copenhaver Et Al
    US 2011 O165635A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2011/0165635 A1 Copenhaver et al. (43) Pub. Date: Jul. 7, 2011 (54) METHODS AND MATERALS FOR Publication Classification PROCESSINGA FEEDSTOCK (51) Int. Cl. CI2P I 7/04 (2006.01) (75) Inventors: Gregory P. Copenhaver, Chapel CI2P I/00 (2006.01) Hill, NC (US); Daphne Preuss, CI2P 7/04 (2006.01) Chicago, IL (US); Jennifer Mach, CI2P 7/16 (2006.01) Chicago, IL (US) CI2P 7/06 (2006.01) CI2P 5/00 (2006.01) CI2P 5/02 (2006.01) (73) Assignee: CHROMATIN, INC., Chicago, IL CI2P3/00 (2006.01) (US) CI2P I/02 (2006.01) CI2N 5/10 (2006.01) (21) Appl. No.: 12/989,038 CI2N L/15 (2006.01) CI2N I/3 (2006.01) (52) U.S. Cl. ........... 435/126; 435/41; 435/157; 435/160; (22) PCT Fled: Apr. 21, 2009 435/161; 435/166; 435/167; 435/168; 435/171; 435/419,435/254.11: 435/257.2 (86) PCT NO.: PCT/US2O09/041260 (57) ABSTRACT S371 (c)(1), The present disclosure relates generally to methods for pro (2), (4) Date: Mar. 11, 2011 cessing a feedstock. Specifically, methods are provided for processing a feedstock by mixing the feedstock with an addi tive organism that comprises one or more transgenes coding Related U.S. Application Data for one or more enzymes. The expressed enzymes may be (60) Provisional application No. 61/046,705, filed on Apr. capable of breaking down cellulosic and lignocellulosic 21, 2008. materials and converting them to a biofuel.
    [Show full text]
  • Enzymatic Production of Human Milk Oligosaccharides
    Downloaded from orbit.dtu.dk on: Oct 10, 2021 Enzymatic production of human milk oligosaccharides Guo, Yao Publication date: 2014 Document Version Publisher's PDF, also known as Version of record Link back to DTU Orbit Citation (APA): Guo, Y. (2014). Enzymatic production of human milk oligosaccharides. DTU Chemical Engineering. General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights. Users may download and print one copy of any publication from the public portal for the purpose of private study or research. You may not further distribute the material or use it for any profit-making activity or commercial gain You may freely distribute the URL identifying the publication in the public portal If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim. Enzymatic production of human milk oligosaccharides Yao Guo Ph.D. Thesis April 2014 Enzymatic production of human milk oligosaccharides Yao Guo Ph.D. thesis, April 2014 Center for BioProcess Engineering 1 Copyright©: Yao Guo April 2014 Center For Bioprocess Engineering Department of Chemical and Biochemical Engineering Technical University of Denmark Søltofts Plads, Building 229 DK-2800 Kgs. Lyngby Denmark Phone: +45 4525 2800 Fax: +45 4525 2258 Web: www.bioeng.kt.dtu.dk Print: J&R Frydenberg A/S København September 2014 ISBN: 978-87-93054-43-1 2 Preface This thesis comprises the research carried out during my PhD study at the Center for BioProcess Engineering, Department of Chemical and Biochemical Engineering, Technical University of Denmark, from August 2010 to April 2014.
    [Show full text]