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  • University of Huddersfield Repository

    Sutcliffe, Victoria

    Synthesis and Reactivity of -Sultams with the Potential to Act as Metallo-Enzyme Inhibitorsβ

    Original Citation

    Sutcliffe, Victoria (2013) Synthesis and Reactivity of -Sultams with the Potential to Act as β

    Metallo-Enzyme Inhibitors. Masters thesis, University of Huddersfield.

    This version is available at http://eprints.hud.ac.uk/19279/

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  • SYNTHESIS AND REACTIVITY OF

    β-SULTAMS WITH THE POTENTIAL TO ACT AS METALLO-ENZYME

    INHIBITORS

    VICTORIA LOUISE SUTCLIFFE

    A thesis submitted to the University of Huddersfield in partial fulfilment of the

    requirements for the degree of Master of Philosophy

    The University of Huddersfield

    Submission date: August 2013

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    Abstract

    Enzyme inhibition forms the basis of much of the medicinal chemistry used in the treatment of disease. -Sultams are cyclic sulfonamides which are both -lactam analogues and potential pro-drugs of taurine and substituted taurines: as their hydrolysis products. Two -sultams, their hydrolysis products and a range of dicarboxylic acids were tested as inhibitors of BcII, a metallo- -lactamase enzyme. The two -sultams, their hydrolysis products and some related compounds were also tested as inhibitors of glutamine synthetase following work showing that -sultam has an effect on neurotransmission in the CNS. A novel -sultam, 1,2-thiazetidine-3-carboxylate-1,1-dioxide (3-carboxy- -sultam), has been synthesised via a four-step process from L-cystine including the removal of a benzyl ester group from benzyl 3-carboxylate- -sultam utilising sodium in liquid ammonia. The product has been characterised by NMR and MS. The rate of hydrolysis of 3-carboxy- -sultam was investigated using 1H NMR and a pH-rate profile produced showing two hydrolysis processes on the acidic limb both of which were first order in hydronium ion concentration (kH = 2.00 x 10

    -1 and 4.8 M-1s-1 respectively) and an alkali catalysed limb first order in hydroxide concentration, kOH = 5.00 x 10

    -4 M-1s-1. The half-life of 3-carboxy- -sultam at physiological pH is approximately 16.5 days. The rate of hydrolysis of the unsubstituted -sultam at acidic pH was investigated by ReactIR and shown to be first order in hydronium ion concentration though kH was not calculated due to variations in the quality of the collected data. Neither the 3-carboxy- -sultam nor the unsubstituted -sultam inhibited BcII or glutamine synthetase. D-Cysteine is a weak inhibitor of BcII, Ki = 7.5 x 10

    -3 M, and a substrate for glutamine synthetase. L-Cysteine is also a substrate for glutamine synthetase and L-cysteic acid is a very weak inhibitor of BcII. The mechanism of BcII catalysed hydrolysis of ertapenem was investigated using 1H NMR and shown to proceed via protonation of the ring opened pyrrolidine ring at C3 leading to the formation of an imine.

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    Table of Contents

    Abstract ........................................................................................................ 3

    Table of Contents ......................................................................................... 4

    Acknowledgements ...................................................................................... 7

    List of abbreviations ..................................................................................... 8

    Introduction ............................................................................................... 11

    1.1 Background and Aims of Research ...................................................................................................................12

    1.2 Bacteria, Antibiotics and Resistance .................................................................................................................14

    1.2.1 Bacteria ......................................................................................................................................................14

    1.2.2 Antibiotics ..................................................................................................................................................17

    1.2.3 Antibiotic Resistance ..................................................................................................................................23

    1.3 β-Lactamase Enzymes .......................................................................................................................................28

    1.4 Active Site Serine β-Lactamases; Classes A, C and D.........................................................................................32

    1.5 Metallo-β-Lactamases; Class B .........................................................................................................................38

    1.5.1 Mechanisms of Metallo-β-Lactamases ........................................................................................................40

    1.5.2 Metallo-β-Lactamase from Bacillus cereus; BcII ..........................................................................................43

    1.6 Inhibition of Class B Metallo-β-Lactamases ......................................................................................................46

    1.7 β-Sultams..........................................................................................................................................................51

    1.7.1 Structure ....................................................................................................................................................51

    1.7.2 Synthesis ....................................................................................................................................................52

    1.7.3 Reactivity....................................................................................................................................................60

    1.7.4 Sulfonyl Compounds as Enzyme Inhibitors ..................................................................................................65

    . . β-Sultams as Enzyme Inhibitors ..................................................................................................................67

    1.8 Glutamine Synthetase ......................................................................................................................................71

    1.8.1 The Roles and Regulation of Glutamic Acid in The Human Body .................................................................71

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    1.8.2 The Role of Glutamic Acid in Neurodegeneration .......................................................................................72

    1.8.3 Glutamic Acid Analogues (and Associated Compounds) as Enzyme Inhibitors and CNS Drugs .....................73

    . . β-Sultams as Pro-Drugs for Treating Neurodegenerative Diseases ..............................................................75

    1.9 Mechanism of Ertapenem Hydrolysis by the Metallo-β-Lactamase enzyme BcII .............................................77

    1.10 Instrumental Techniques – React-IR ..............................................................................................................79

    Experimental .................................................