Crystal Structure Analysis and in Silico pKa Calculations Suggest Strong pKa Shifts of Ligands as Driving Force for High‐Affinity Binding to TGT

Expanded lin‐benzoguanines exhibit binding affinities to tRNA‐guanine transglycosylase (TGT) in the low‐nanomolar range. A significant pKa shift is observed for the inhibitors moving from aqueous solution to protein environment. The protonation of the inhibitor facilitates a charge‐assisted hydrogen bond in the protein–ligand complex.

[1]  J. Gasteiger,et al.  ITERATIVE PARTIAL EQUALIZATION OF ORBITAL ELECTRONEGATIVITY – A RAPID ACCESS TO ATOMIC CHARGES , 1980 .

[2]  G. Klebe,et al.  Crystal structures of tRNA-guanine transglycosylase (TGT) in complex with novel and potent inhibitors unravel pronounced induced-fit adaptations and suggest dimer formation upon substrate binding. , 2007, Journal of molecular biology.

[3]  D. Suck,et al.  Purification, crystallization, and preliminary X‐ray diffraction studies of tRNA‐guanine transglycosylase from Zymomonas mobilis , 1996, Proteins.

[4]  K. Koch,et al.  tRNA-guanine transglycosylase from Escherichia coli. Overexpression, purification and quaternary structure. , 1993, Journal of molecular biology.

[5]  G. Björk,et al.  Transfer RNA modification, temperature and DNA superhelicity have a common target in the regulatory network of the virulence of Shigella flexneri: the expression of the virF gene , 2000, Molecular microbiology.

[6]  S. Nishimura,et al.  Isolation and characterization of a guanine insertion enzyme, a specific tRNA transglycosylase, from Escherichia coli. , 1979, The Journal of biological chemistry.

[7]  Z. Otwinowski,et al.  [20] Processing of X-ray diffraction data collected in oscillation mode. , 1997, Methods in enzymology.

[8]  J. Richardson,et al.  Asparagine and glutamine: using hydrogen atom contacts in the choice of side-chain amide orientation. , 1999, Journal of molecular biology.

[9]  J. Thornton,et al.  PROCHECK: a program to check the stereochemical quality of protein structures , 1993 .

[10]  A. Hopkins,et al.  Ligand efficiency: a useful metric for lead selection. , 2004, Drug discovery today.

[11]  G. Klebe,et al.  Mechanism and Substrate Specificity of tRNA–Guanine Transglycosylases (TGTs): tRNA‐Modifying Enzymes from the Three Different Kingdoms of Life Share a Common Catalytic Mechanism , 2005, Chembiochem : a European journal of chemical biology.

[12]  M. Watarai,et al.  vacC, a virulence-associated chromosomal locus of Shigella flexneri, is homologous to tgt, a gene encoding tRNA-guanine transglycosylase (Tgt) of Escherichia coli K-12 , 1994, Journal of bacteriology.

[13]  G Klebe,et al.  A new target for shigellosis: rational design and crystallographic studies of inhibitors of tRNA-guanine transglycosylase. , 2000, Journal of molecular biology.

[14]  Gerhard Klebe,et al.  Development, validation, and application of adapted PEOE charges to estimate pKa values of functional groups in protein–ligand complexes , 2006, Proteins.

[15]  G. Klebe,et al.  Glutamate versus glutamine exchange swaps substrate selectivity in tRNA-guanine transglycosylase: insight into the regulation of substrate selectivity by kinetic and crystallographic studies. , 2007, Journal of molecular biology.

[16]  Gerhard Klebe,et al.  Protonation changes upon ligand binding to trypsin and thrombin: structural interpretation based on pK(a) calculations and ITC experiments. , 2007, Journal of molecular biology.

[17]  Kevin Cowtan,et al.  research papers Acta Crystallographica Section D Biological , 2005 .

[18]  W. Xie,et al.  Chemical trapping and crystal structure of a catalytic tRNA guanine transglycosylase covalent intermediate , 2003, Nature Structural Biology.

[19]  D. Suck,et al.  Crystal structure of tRNA‐guanine transglycosylase: RNA modification by base exchange. , 1996, The EMBO journal.

[20]  D. M. Goodenough-Lashua,et al.  tRNA-guanine transglycosylase from E. coli: a ping-pong kinetic mechanism is consistent with nucleophilic catalysis. , 2003, Bioorganic chemistry.

[21]  A. Curnow,et al.  tRNA-guanine Transglycosylase from Escherichia coli , 1995, The Journal of Biological Chemistry.

[22]  G. Klebe,et al.  Hochaffine Inhibitoren der tRNA‐Guanin‐Transglycosylase, eines Schlüsselenzyms in der Pathogenese der Shigellen‐Ruhr: ladungsverstärkte Wasserstoffbrücken , 2007 .

[23]  K. Watanabe,et al.  A UGU sequence in the anticodon loop is a minimum requirement for recognition by Escherichia coli tRNA-guanine transglycosylase. , 1994, The Journal of biological chemistry.

[24]  G. Klebe,et al.  Potent inhibitors of tRNA-guanine transglycosylase, an enzyme linked to the pathogenicity of the Shigella bacterium: charge-assisted hydrogen bonding. , 2007, Angewandte Chemie.

[25]  K. Reuter,et al.  Sequence analysis and overexpression of the Zymomonas mobilis tgt gene encoding tRNA-guanine transglycosylase: purification and biochemical characterization of the enzyme , 1995, Journal of bacteriology.

[26]  R J Read,et al.  Crystallography & NMR system: A new software suite for macromolecular structure determination. , 1998, Acta crystallographica. Section D, Biological crystallography.

[27]  J. L. Smith,et al.  Structure of myohemerythrin in the azidomet state at 1.7/1.3 A resolution. , 1987, Journal of molecular biology.

[28]  G. Klebe,et al.  Synthesis, Biological Evaluation, and Crystallographic Studies of Extended Guanine-Based (lin-Benzoguanine) Inhibitors for tRNA-Guanine Transglycosylase (TGT) , 2006 .

[29]  G. Sheldrick,et al.  SHELXL: high-resolution refinement. , 1997, Methods in enzymology.

[30]  G. Klebe,et al.  Thermodynamisches Inhibitionsprofil eines Cyclopentyl- und eines Cyclohexylderivats gegenüber Thrombin: gleich, jedoch aus unterschiedlichem Grund† , 2007 .

[31]  Alex M. Clark,et al.  2D Structure Depiction , 2006, J. Chem. Inf. Model..

[32]  Z. Samra,et al.  Growing antimicrobial resistance of Shigella isolates. , 2003, The Journal of antimicrobial chemotherapy.

[33]  Gerhard Klebe,et al.  Thermodynamic inhibition profile of a cyclopentyl and a cyclohexyl derivative towards thrombin: the same but for different reasons. , 2007, Angewandte Chemie.

[34]  P Willett,et al.  Development and validation of a genetic algorithm for flexible docking. , 1997, Journal of molecular biology.

[35]  Gerhard Klebe,et al.  Atypical Protonation States in the Active Site of HIV-1 Protease: A Computational Study , 2007, J. Chem. Inf. Model..

[36]  W. Delano.  The PyMOL Molecular Graphics System , 2002 .