From lin-benzoguanines to lin-benzohypoxanthines as ligands for Zymomonas mobilis tRNA-guanine transglycosylase: replacement of protein-ligand hydrogen bonding by importing water clusters.
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, NH(2), or NH-alkyl. These differences lead to major changes in the inhibition constants, pK(a) values, and binding modes. Compared to the lin-benzoguanines, with an exocyclic NH(2) at C(6), the lin-benzohypoxanthines without an exocyclic NH(2) 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 NH(2) 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.
Laboratorium für Organische Chemie, ETH Zürich, Hönggerberg, HCI, 8093 Zurich, Switzerland.
Luzi Jakob
LJ
Barandun
Florian
F
Immekus
Philipp C
PC
Kohler
Sandro
S
Tonazzi
Björn
B
Wagner
Severin
S
Wendelspiess
Tina
T
Ritschel
Andreas
A
Heine
Manfred
M
Kansy
Gerhard
G
Klebe
François
F
Diederich
D001665:Binding Sites; D018360:Crystallography, X-Ray; D004405:Dysentery, Bacillary; D006147:Guanine / Q000031:analogs & derivatives* / Q000737:chemistry*; D006860:Hydrogen Bonding; D019271:Hypoxanthine / Q000737:chemistry*; D008024:Ligands; D008958:Models, Molecular; D015394:Molecular Structure; D010430:Pentosyltransferases / Q000737:chemistry*; D011485:Protein Binding; D012763:Shigella flexneri / Q000737:chemistry* / Q000201:enzymology*; D014867:Water / Q000737:chemistry*; D017355:Zymomonas / Q000737:chemistry* / Q000201:enzymology*
D008024:Ligands; D014867:Water; D019271:Hypoxanthine; D006147:Guanine; D010430:Pentosyltransferases; C020837:queuine tRNA-ribosyltransferase