From lin-Benzoguanines to lin-Benzohypoxanthines as Ligands for Zymomonas mobilis tRNA–Guanine Transglycosylase: Replacement of Protein–Ligand Hydrogen Bonding by Importing Water Clusters

Authors

  • Luzi Jakob Barandun,

    1. Laboratorium für Organische Chemie, ETH Zürich, Hönggerberg, HCI, 8093 Zurich (Switzerland), Fax: (+41) 44-632-1109
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    • These authors contributed equally to this work.

  • Florian Immekus,

    1. Institut für Pharmazeutische Chemie, Philipps-Universität Marburg, Marbacher Weg 6, 35032 Marburg (Germany), Fax: (+49) 6421-282-8994
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    • These authors contributed equally to this work.

  • Dr. Philipp C. Kohler,

    1. Laboratorium für Organische Chemie, ETH Zürich, Hönggerberg, HCI, 8093 Zurich (Switzerland), Fax: (+41) 44-632-1109
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  • Sandro Tonazzi,

    1. Laboratorium für Organische Chemie, ETH Zürich, Hönggerberg, HCI, 8093 Zurich (Switzerland), Fax: (+41) 44-632-1109
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  • Björn Wagner,

    1. Pharma Research Non-Clinical Safety, F. Hoffmann-La Roche AG, 4070 Basel (Switzerland)
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  • Severin Wendelspiess,

    1. Pharma Research Non-Clinical Safety, F. Hoffmann-La Roche AG, 4070 Basel (Switzerland)
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  • Dr. Tina Ritschel,

    1. Institut für Pharmazeutische Chemie, Philipps-Universität Marburg, Marbacher Weg 6, 35032 Marburg (Germany), Fax: (+49) 6421-282-8994
    2. Current address: Computational Drug Discovery Group, CMBI, Radboud University Nijmegen Medical Centre, P.O. Box 9101, 6500 HB Nijmegen (The Netherlands)
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  • Dr. Andreas Heine,

    1. Institut für Pharmazeutische Chemie, Philipps-Universität Marburg, Marbacher Weg 6, 35032 Marburg (Germany), Fax: (+49) 6421-282-8994
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  • Dr. Manfred Kansy,

    1. Pharma Research Non-Clinical Safety, F. Hoffmann-La Roche AG, 4070 Basel (Switzerland)
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  • Prof. Dr. Gerhard Klebe,

    Corresponding author
    1. Institut für Pharmazeutische Chemie, Philipps-Universität Marburg, Marbacher Weg 6, 35032 Marburg (Germany), Fax: (+49) 6421-282-8994
    • Institut für Pharmazeutische Chemie, Philipps-Universität Marburg, Marbacher Weg 6, 35032 Marburg (Germany), Fax: (+49) 6421-282-8994
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  • Prof. Dr. François Diederich

    Corresponding author
    1. Laboratorium für Organische Chemie, ETH Zürich, Hönggerberg, HCI, 8093 Zurich (Switzerland), Fax: (+41) 44-632-1109
    • Laboratorium für Organische Chemie, ETH Zürich, Hönggerberg, HCI, 8093 Zurich (Switzerland), Fax: (+41) 44-632-1109
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Abstract

The foodborne illness shigellosis is caused by Shigella bacteria that secrete the highly cytotoxic Shiga toxin, which is also formed by the closely related enterohemorrhagic Escherichia coli (EHEC). It has been shown that tRNA–guanine transglycosylase (TGT) is essential for the pathogenicity of Shigella flexneri. Herein, the molecular recognition properties of a guanine binding pocket in Zymomonas mobilis TGT are investigated with a series of lin-benzohypoxanthine- and lin-benzoguanine-based inhibitors that bear substituents to occupy either the ribose-33 or the ribose-34 pocket. The three inhibitor scaffolds differ by the substituent at C(6) being H, NH2, or NH[BOND]alkyl. These differences lead to major changes in the inhibition constants, pKa values, and binding modes. Compared to the lin-benzoguanines, with an exocyclic NH2 at C(6), the lin-benzohypoxanthines without an exocyclic NH2 group have a weaker affinity as several ionic protein–ligand hydrogen bonds are lost. X-ray cocrystal structure analysis reveals that a new water cluster is imported into the space vacated by the lacking NH2 group and by a conformational shift of the side chain of catalytic Asp102. In the presence of an N-alkyl group at C(6) in lin-benzoguanine ligands, this water cluster is largely maintained but replacement of one of the water molecules in the cluster leads to a substantial loss in binding affinity. This study provides new insight into the role of water clusters at enzyme active sites and their challenging substitution by ligand parts, a topic of general interest in contemporary structure-based drug design.

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