Assembling and Unraveling Carbohydrates Structures 957006/  · Assembling and Unraveling

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  • Assembling and Unraveling Carbohydrates Structures Conformational analysis of synthesized branched oligosaccharides

    Thibault Angles dOrtoli

  • Thibault Angles dOrtoli, Stockholm University 2016 Cover picture: Sugar Tetris ISBN 978-91-7649-473-8 Printed by Holmbergs, Malm 2016 Distributor: Department of Organic Chemistry

  • I am finetuning my soul To the universal wavelength No one is a lover alone I propose an atom dance. Bjrk & Anohni Atom Dance

  • - Till Rasmus

  • v

    Abstract

    Advances in the elaboration of vaccines and enzyme inhibitors rely on acquir-ing more knowledge about protein-carbohydrate binding events. The relation-ships between biological function and the three-dimensional properties of large glycans can be studied by focusing on the structural components they contain, namely, by scaling down the system under analysis. Chemical meth-ods are useful assets as they allow the determination and isolation of epitopes; these small and recognizable fragments that lead to very specific interactions. In this thesis, biologically relevant saccharides were obtained using recently developed concepts in carbohydrate synthesis and NMR spectroscopy was used to unravel their conformational preferences.

    In paper I, the convergent synthesis of the tetrasaccharide found in the nat-ural product solaradixine is described. Reactivity enhanced disaccharide gly-cosyl donors were coupled to a disaccharide acceptor in a 2 + 2 fashion. The computer program CASPER was subsequently used to verify the synthesized structure.

    The conformation arming concept employed in paper I was further investi-gated in paper II. An NMR-based methodology enabled the determination of the ring conformations of a set of donors. Subsequenttly, glycosylation reac-tions were performed and yields were correlated to donors ring shapes. Per-turbations in the rings shape caused by bulky silyl ether protective groups were sufficient to boost the potency of several donors. As a matter of fact, complex branched oligosaccharides could be obtained in good to excellent yields.

    In paper III, NMR spectroscopy observables were measured to elucidate the ring shape, the mutual orientation of the rings across the glycosidic bond and the positions of the side chains of 5 trisaccharides found in larger struc-tures. With the aid of molecular dynamics simulations, their overall confor-mational propensities were revealed.

    Finally, the prediction skills of the software CASPER were improved by adding, inter alia, NMR information of synthesized mono- and disaccharides to its database. Unassigned chemical shifts from a branched heptasaccharide repeating unit served as input to challenge its ability to solve large carbohy-drate structures.

  • vi

  • vii

    List of Publications

    This thesis is based on the following papers, which will be referred to by Roman numerals I IV. Reprints were made with the kind permission of the pusblishers.

    I. Synthesis of the Tetrasaccharide Glycoside Moiety of Solaradix-ine and Rapid NMR-based Structure Verification Using the Pro-gram CASPER Angles dOrtoli, T.; Widmalm, G. Tetrahedron. 2016, 72, 912 927. Supporting information available online

    II. Structure-Reactivity Relationships of Conformationally Armed Disaccharide Donors and Their Use in Synthesis of a Hexasaccha-ride Related to the Capsular Polysaccharide from Streptococcus pneumoniae type 37 Angles dOrtoli, T.; Hamark, C.; Widmalm, G. In manuscript.

    III. Delineating the Conformational Flexibility of Trisaccharides from NMR Spectroscopy Experiments and Computer Simula-tions Yang, M.; Angles dOrtoli, T.; Swn, E.; Jana, M.; Widmalm, G.; MacKerell, Jr., A. D. Phys. Chem. Chem. Phys.. 2016, 18, 18776 18794. Supporting information available online

    IV. Complete 1H and 13C NMR Chemical Shift Assignments of Mono-

    to Tetrasaccharides as Basis for NMR Chemical Shift Predictions of Oligo- and Polysaccharides Using the Computer Program CASPER Angles dOrtoli, T.; Mobarak, H.; Sthle, J.; Hamark, C.; Fontana, C.; Engstrm, O.; Apostolica, P.; Widmalm, G. In manuscript.

  • viii

    Also published: Complete 1H and 13C NMR Chemical Shift Assignments of Mono- to Tetrasaccharides as Basis for NMR Chemical Shift Predictions of Oligosaccharides Using the Computer Program CASPER Rnnols, J.; Pendrill, P.; Fontana, C.; Hamark, C.; Angles dOr-toli, T.; Engstrm, O.; Sthle, J.; Zaccheus, M. V.; Swn, E.; Hahn, L. E.; Iqbal, S.; Widmalm, G. Carb. Res. 2013, 380, 156 166. Temperature Dependence of Hydroxymethyl Group Rota-mer Populations in Cellooligomers Angles dOrtoli, T.; Sjoberg, N. A.; Vasiljeva, P.; Lindman, J.; Widmalm, G.; Bergenstrahle-Wohlert, M.; Wohlert, J. J. Phys. Chem. B. 2015, 119, 9559 9570.

    On the Anomalous Temperature Dependence of Cellulose Aqueous Solubility Bergenstrahle-Wohlert, M.; Angles dOrtoli, T.; Sjoberg, N. A.; Widmalm, G.; Wohlert, J. Cellulose. 2016, 23, 2375 2387. A Molecular Dynamics Study of the Effect of Glycosidic Linkage Type in the Hemicellulose Backbone on the Molec-ular Chain Flexibility Berglund, J.; Angles d'Ortoli, T.; Vilaplana, F.; Widmalm, G.; Bergenstrhle-Wohlert, M.; Lawoko, M.; Henriksson, G.; Lindstrm, M.; Wohlert, J. Accepted in: The Plant Journal.

  • ix

    Contents

    1 ........................................................................................................... 1-iv

    Abstract ......................................................................................................... v

    List of Publications .................................................................................... vii

    Abbreviations ............................................................................................... xi

    1 Introduction ......................................................................................... 1 1.1 Introduction to carbohydrates ...................................................................... 2 1.2 Structure of carbohydrates ........................................................................... 4 1.3 Conformation of carbohydrates .................................................................... 6

    2 Synthesis of Carbohydrates ........................................................... 10 2.1 Enzymatic synthesis ..................................................................................... 10 2.2 Chemical synthesis ....................................................................................... 12

    3 Analytical Methods ........................................................................... 17 3.1 NMR spectroscopy ........................................................................................ 17

    3.1.1 Chemical shifts ................................................................................... 18 3.1.2 Coupling constants............................................................................. 19 3.1.3 The nuclear Overhauser effect ........................................................ 23

    3.2 Molecular dynamics simulations in conformational analysis ................. 25

    4 Synthesis of the Tetrasaccharide Glycoside Moiety of Solaradixine and Rapid NMR-Based Structure Verification Using the Program CASPER (Paper I)...................................................................... 26

    4.1 Synthesis ........................................................................................................ 28 4.1.1 Synthesis of monosaccharide acceptor 8 ...................................... 28 4.1.2 Synthesis of disaccharide 14 ........................................................... 29 4.1.3 Synthesis of trisaccharide 20 .......................................................... 30 4.1.4 Synthesis of tetrasaccharide 1 ........................................................ 32

    4.2 Characterization of tetrasaccharide 1 by NMR spectroscopy ................ 35 4.3 The program CASPER as a tool for fast structure verification .............. 36 4.4 Conclusion ...................................................................................................... 37

    5 Structure-Reactivity Relationships of Conformationally Armed Disaccharide Donors and Their Use in Synthesis of a Hexasaccharide

  • x

    Related to the Capsular Polysaccharide from Streptococcus pneumoniae type 37 (Paper II) .............................................................. 39

    5.1 Glycosyl donors ............................................................................................. 40 5.2 Trisaccharides and tetrasaccharides ......................................................... 42 5.3 Hexasaccharide ............................................................................................. 47 5.4 Conclusion ...................................................................................................... 51

    6 Delineating the Conformational Flexibility of Trisaccharides from NMR Spectroscopy Experiments and Computer Simulations (Paper III) ................................................................................................... 52

    6.1 Synthesis ........................................................................................................ 54 6.2 NMR spectroscopy ........................................................................................ 55

    6.2.1 1H,1H internuclear distances ............................................................ 55 6.2.2 Spin-spin coupling constants ........................................................... 57 6.2.3 Comparison between MD and NMR spectroscopy results ........... 58 6.2.4 Conformational preferences of the trisaccharides ........................ 60

    6.3 Con