A Practical Enzymatic Synthesis of UDP Sugars and NDP Glucoses
2005; Wiley; Volume: 6; Issue: 11 Linguagem: Inglês
10.1002/cbic.200500183
ISSN1439-7633
AutoresJungdon Bae, Kwang‐Ho Kim, Doo‐Il Kim, Yongseok Choi, Joong Su Kim, Sukhoon Koh, Suk‐In Hong, Dae‐Sil Lee,
Tópico(s)Enzyme Production and Characterization
ResumoChemBioChemVolume 6, Issue 11 p. 1963-1966 Communication A Practical Enzymatic Synthesis of UDP Sugars and NDP Glucoses Jungdon Bae Dr., Jungdon Bae Dr. Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597 School of Life sciences and Biotechnology, Korea University, Seoul 136-701, KoreaSearch for more papers by this authorKwang-Ho Kim, Kwang-Ho Kim Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597Search for more papers by this authorDooil Kim, Dooil Kim Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597 Department of Biomicrosystem Technology, Korea University, Seoul 136-701, KoreaSearch for more papers by this authorYongseok Choi Dr., Yongseok Choi Dr. Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597Search for more papers by this authorJoong Su Kim Dr., Joong Su Kim Dr. Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597Search for more papers by this authorSukhoon Koh Dr., Sukhoon Koh Dr. Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597Search for more papers by this authorSuk-In Hong Prof., Suk-In Hong Prof. Department of Chemical and Biological Engineering, Korea University, Seoul 136-701, KoreaSearch for more papers by this authorDae-Sil Lee Dr., Dae-Sil Lee Dr. [email protected] Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597Search for more papers by this author Jungdon Bae Dr., Jungdon Bae Dr. Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597 School of Life sciences and Biotechnology, Korea University, Seoul 136-701, KoreaSearch for more papers by this authorKwang-Ho Kim, Kwang-Ho Kim Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597Search for more papers by this authorDooil Kim, Dooil Kim Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597 Department of Biomicrosystem Technology, Korea University, Seoul 136-701, KoreaSearch for more papers by this authorYongseok Choi Dr., Yongseok Choi Dr. Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597Search for more papers by this authorJoong Su Kim Dr., Joong Su Kim Dr. Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597Search for more papers by this authorSukhoon Koh Dr., Sukhoon Koh Dr. Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597Search for more papers by this authorSuk-In Hong Prof., Suk-In Hong Prof. Department of Chemical and Biological Engineering, Korea University, Seoul 136-701, KoreaSearch for more papers by this authorDae-Sil Lee Dr., Dae-Sil Lee Dr. [email protected] Genome Research Center, Korea Research Institute of Bioscience and Biotechnology, Yuseong, Daejeon 305-333, Korea, Fax: (+82) 42-860-4597Search for more papers by this author First published: 26 October 2005 https://doi.org/10.1002/cbic.200500183Citations: 23Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onEmailFacebookTwitterLinkedInRedditWechat Graphical Abstract Mix and match. Nucleotide sugars are essential as glycoside donors to glycosyltransferases for the synthesis of bioactive metabolites. With 45 combinations of reactions with five NTPs and nine sugar-1-phosphates (Su1Ps) by recombinant UDP-sugar pyrophosphorylase (UP) from Thermus caldophilus GK24, the enzyme showed broad substrate specificity toward five types of NTP with glucose-1-phosphate as well as four Su1Ps with UTP, affording all nine nucleotide sugars. Supporting Information Supporting information for this article is available on the WWW under http://www.wiley-vch.de/contents/jc_2268/2005/z500183_s.pdf or from the author. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article. References 1 1aL. Elling, Adv. Biochem. Eng./Biotechnol. 1997, 58, 89; 1bT. Bülter, L. Elling, Glycoconjugate J. 1999, 16, 147. 2 2aY. Ichikawa, G. C. Look, C. H. Wong, Anal. Biochem. 1992, 202, 215; 2bD. Maskell, C. R. Raetz, P. D. Rick, Trends Microbiol. 1996, 4, 495; 2cM. M. Palcic, Curr. Opin. Biotechnol. 1999, 10, 616. 3 3aJ. H. Ko, C. H. Kim, D. S. Lee, Y. S. Kim, Biochem. J. 1996, 319, 977; 3bN. Parajuli, D. S. Lee, H. C. Lee, K. Liou, J. K. Sohng, Biotechnol. Lett. 2004, 26, 437. 4 4aJ. S. Thorson, W. A. Barton, D. Hoffmeister, C. Albermann, D. B. Nikolov, ChemBioChem 2004, 5, 16; 4bJ. Yang, D. Hoffmeister, L. Liu, X. Fu, J. S. Thorson, Bioorg. Med. Chem. 2004, 12, 1577; 4cJ. S. Thorson, E. L. Sievers, J. Ahlert, E. Shepard, R. E. Whitwam, K. C. Onwueme, M. Ruppen, Curr. Pharm. Des. 2000, 6, 1841. 5 5aS. Roseman, J. J. Distler, J. G. Moffatt, H. G. Khorana, J. Am. Chem. Soc. 1961, 83, 659; 5bD. M. Carlson, R. G. Hansen, J. Biol. Chem. 1962, 237, 1260; 5cJ. E. Heidlas, W. J. Lees, P. Pale, G. M. 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Zea, N. L. Pohl, J. Am. Chem. Soc. 2004, 126, 15993; 8cT. Kotake, D. Yamaguchi, H. Ohzono, S. Hojo, S. Kaneko, H. K. Ishida, Y. Tsumuraya, J. Biol. Chem. 2004, 279, 45728. 9The Tca usp gene was obtained from the genome data by shot-gun sequencing, and compared with the reported 15 N-terminal amino acid residues (MKGLILAAGRGTRLR) of purified UP, which was amplified from Tca genomic DNA by a polymerase chain reaction. 10The transformed cells, harvested by centrifugation, were resuspended in buffer A (25 mL, 20 mM Tris-HCl, pH 7.0, 1 mM β-mercaptoethanol), disrupted by sonication, and centrifuged. The supernatant was heat-precipitated at 80 °C for 20 min and filtered, then loaded onto a DEAE-Sephacel column (2×20 cm—after equilibration with buffer A, the enzyme was eluted by a linear gradient of 0–0.3 M KCl at 1 mL min−1 on a FPLC system), and pooled, concentrated, and dialyzed with buffer A. 11Glc1P, glucose-1-phosphate; Gal1P, galactose-1-phosphate; GlcNAc1P, N-acetylglucosamine-1-phosphate; Man1P, mannose-1-phosphate; Xyl1P, xylose-1-phosphate, GlcN1P, glucosamine-1-phosphate; Fuc1P, fucose-1-phosphate; Fru1P, fructose-1-phosphate; Mal1P, maltose-1-phosphate. 12The standard assay mixture, containing Tris-HCl (50 mM, pH 8.0), MgCl2 (8 mM), Su1P (5 mM), and NTP (5 mM), was incubated at 70 °C for 10 min in a final volume of 50 μL. One unit of the enzyme is defined as the amount of enzyme catalyzing the formation of 1 μmol of UDP-GlcNAc per min at 70 °C. 13The HPLC conditions were described with sight modification in ref. [8a]. Assays of the recombinant Tca UP were measured by the amount of NDP sugar formed from NTP and Su1P by using HPLC (Beckman Instruments, Palo Alto, CA). A Hypersil ODS C18 reversed-phase column was used (3 μm, 4.6×250 mm; Phenomenex, Torra, CA), with tetrabutylammonium chloride as ion-pair reagent. The column was equilibrated with 90 % (v/v) solvent A (12.2 mM tetrabutylammonium chloride in 10.2 mM KH2PO4, 5 % acetonitrile, pH 4.0) and 10 % (v/v) solvent B (70 % (v/v) acetonitrile in solvent A). The reaction products were eluted on the column with a gradient of solvent B in solvent A (10–90 % solvent B for 15 min, then 90–100 % solvent B for 1 min). Standard samples (NMP, NDP, NTP, and NDP sugars) were prepared at concentrations of 0.02–0.5 mM. Peaks were identified from their retention times, obtained from the absorbance at 254 nm at 1 mL min−1. 14J. H. Ko, C. H. Kim, D. S. Lee, Y. S. Kim, Biochem. J. 1996, 319, 977. 15A. Silipo, A. Molinaro, C. De Castro, R. Ferrara, I. Romano, B. Nicolaus, R. Lanzetta, M. Parrilli, Eur. J. org. Chem. 2004, 5047. 16Z. Silva, S. Alarico, M. S. da Costa, Extremophiles 2005, 9, 29. 17 17aK. Brown, F. Pompeo, S. Dixon, D. Mengin-Lecreulx, C. Cambillau, Y. Bourne, EMBO J. 1999, 18, 4096; 17bW. Blankenfeldt, M. Asuncion, J. S. Lam, J. H. Naismith, EMBO J. 2000, 19, 6652; 17cG. Sulzenbacher, L. Gal, C. Peneff, F. Fassy, Y. Bourne, J. Biol. Chem. 2001, 276, 11844; 17dD. Kostrewa, A. D′Arcy, B. Takacs, M. Kamber, J. Mol. Biol. 2001, 305, 279; 17eS. Zuccotti, D. Zanardi, C. Rosano, L. Sturla, M. Tonetti, M. Bolognesi, J. Mol. Biol. 2001, 313, 831. Citing Literature Volume6, Issue11November 4, 2005Pages 1963-1966 ReferencesRelatedInformation
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