WaterLOGSY as a method for primary NMR screening: Practical aspects and range of applicability

WaterLOGSY represents a powerful method for primary NMR screening in the identification of compounds interacting with macromolecules, including proteins and DNA or RNA fragments. Several relay pathways are used constructively in the experiment for transferring bulk water magnetization to the ligand. The method is particularly useful for the identification of novel scaffolds of micromolar affinity that can be then optimized using directed screening, combinatorial chemistry, medicinal chemistry and structure-based drug design. The practical aspects and range of applicability of the WaterLOGSY experiment are analyzed in detail here. Competition binding and titration WaterLOGSY permit, after proper correction, the evaluation of the dissociation binding constant. The high sensitivity of the technique in combination with the easy deconvolution of the mixtures for the identification of the active components, significantly reduces the amount of material and time needed for the NMR screening process.

[1]  Jens Klein,et al.  DETECTING BINDING AFFINITY TO IMMOBILIZED RECEPTOR PROTEINS IN COMPOUND LIBRARIES BY HR-MAS STD NMR , 1999 .

[2]  G. Otting,et al.  Protein Hydration Viewed by High-Resolution NMR Spectroscopy: Implications for Magnetic Resonance Image Contrast , 1995 .

[3]  Michael J. Shapiro,et al.  NOE Pumping: A Novel NMR Technique for Identification of Compounds with Binding Affinity to Macromolecules , 1998 .

[4]  A M Gronenborn,et al.  Demonstration of positionally disordered water within a protein hydrophobic cavity by NMR. , 1995, Science.

[5]  Gottfried Otting,et al.  Proton exchange rates from amino acid side chains— implications for image contrast , 1996, Magnetic resonance in medicine.

[6]  C. Dalvit,et al.  Half-filter experiments for assignment, structure determination and hydration analysis of unlabelled ligands bound to 13C/15N labelled proteins , 1999, Journal of biomolecular NMR.

[7]  G. Bodenhausen,et al.  Principles of nuclear magnetic resonance in one and two dimensions , 1987 .

[8]  C. Lepre,et al.  Library design for NMR-based screening. , 2001, Drug discovery today.

[9]  B. Halle,et al.  Residence times of the buried water molecules in bovine pancreatic trypsin inhibitor and its G36S mutant. , 1995, Biochemistry.

[10]  B. Halle,et al.  Water molecules in the binding cavity of intestinal fatty acid binding protein: dynamic characterization by water 17O and 2H magnetic relaxation dispersion. , 1999, Journal of molecular biology.

[11]  Claudio Dalvit,et al.  Efficient multiple-solvent suppression for the study of the interactions of organic solvents with biomolecules , 1998 .

[12]  Michael J. Shapiro,et al.  NOE Pumping. 2. A High-Throughput Method To Determine Compounds with Binding Affinity to Macromolecules by NMR , 2000 .

[13]  R. Kaptein,et al.  Band-selective editing of exchange-relay in protein-water NOE experiments , 1999, Journal of biomolecular NMR.

[14]  Ajay,et al.  The SHAPES strategy: an NMR-based approach for lead generation in drug discovery. , 1999, Chemistry & biology.

[15]  K. Wüthrich.  NMR of proteins and nucleic acids , 1988 .

[16]  M. Shapiro,et al.  Screening Mixtures by Affinity NMR , 1997 .

[17]  B. Meyer,et al.  Screening mixtures for biological activity by NMR. , 1997, European journal of biochemistry.

[18]  Wolfgang Jahnke,et al.  Second-Site NMR Screening with a Spin-Labeled First Ligand , 2000 .

[19]  J. Moore,et al.  NMR screening in drug discovery. , 1999, Current opinion in biotechnology.

[20]  Mengfen Lin,et al.  Diffusion-Edited NMR−Affinity NMR for Direct Observation of Molecular Interactions , 1997 .

[21]  Y. Cheng,et al.  Relationship between the inhibition constant (K1) and the concentration of inhibitor which causes 50 per cent inhibition (I50) of an enzymatic reaction. , 1973, Biochemical pharmacology.

[22]  A. J. Shaka,et al.  One-Dimensional NOE Experiments Using Pulsed Field Gradients , 1997 .

[23]  W. Jahnke,et al.  Spin label enhanced NMR screening. , 2001, Journal of the American Chemical Society.

[24]  G. Otting,et al.  NMR identification of hydrophobic cavities with ow water occupancies in protein structures using small gas molecules , 1997, Nature Structural Biology.

[25]  S. Grzesiek,et al.  The Importance of Not Saturating H2o in Protein NMR : application to Sensitivity Enhancement and Noe Measurements , 1993 .

[26]  Thomas Peters,et al.  Bioaffinity NMR Spectroscopy: Identification of an E‐Selectin Antagonist in a Substance Mixture by Transfer NOE , 1999 .

[27]  J. Oncley,et al.  The specific binding of L-tryptophan to serum albumin. , 1958, The Journal of biological chemistry.

[28]  Dalvit.  Homonuclear 1D and 2D NMR Experiments for the Observation of Solvent-Solute Interactions , 1996, Journal of magnetic resonance. Series B.

[29]  B. Halle,et al.  Protein hydration dynamics in aqueous solution: a comparison of bovine pancreatic trypsin inhibitor and ubiquitin by oxygen-17 spin relaxation dispersion. , 1995, Journal of molecular biology.

[30]  B. Meyer,et al.  Group epitope mapping by saturation transfer difference NMR to identify segments of a ligand in direct contact with a protein receptor. , 2001, Journal of the American Chemical Society.

[31]  P. Hajduk,et al.  Discovering High-Affinity Ligands for Proteins: SAR by NMR , 1996, Science.

[32]  G. Otting.  Erratum to “NMR studies of water bound to biological molecules” , 1998 .

[33]  K Wüthrich,et al.  Protein hydration in aqueous solution. , 1991, Science.

[34]  Bernd Meyer,et al.  Characterization of Ligand Binding by Saturation Transfer Difference NMR Spectroscopy. , 1999, Angewandte Chemie.

[35]  A. Redfield,et al.  Cross relaxation and spin diffusion effects on the proton NMR of biopolymers in H2O. Solvent saturation and chemical exchange in superoxide dismutase. , 1978, FEBS letters.

[36]  Philip M. Dean,et al.  Hydration in drug design. 1. Multiple hydrogen-bonding features of water molecules in mediating protein-ligand interactions , 1995, J. Comput. Aided Mol. Des..

[37]  Stephen W. Fesik,et al.  One-Dimensional Relaxation- and Diffusion-Edited NMR Methods for Screening Compounds That Bind to Macromolecules , 1997 .

[38]  J. Feeney,et al.  The effects of intermediate exchange processes on the estimation of equilibrium constants by NMR , 1979 .

[39]  M. Sundström,et al.  Identification of compounds with binding affinity to proteins via magnetization transfer from bulk water* , 2000, Journal of biomolecular NMR.

[40]  G Schreiber,et al.  Electrostatic enhancement of diffusion-controlled protein-protein association: comparison of theory and experiment on barnase and barstar. , 1998, Journal of molecular biology.

[41]  Jean M. Severin,et al.  Discovery of Potent Nonpeptide Inhibitors of Stromelysin Using SAR by NMR , 1997 .

[42]  H. Berendsen,et al.  Proton magnetic relaxation and spin diffusion in proteins , 1976 .

[43]  G. Otting,et al.  Studies of protein hydration in aqueous solution by direct NMR observation of individual protein-bound water molecules , 1989 .

[44]  R. Kaptein,et al.  Editing of chemical exchange-relayed NOEs in NMR experiments for the observation of protein-water interactions. , 1999, Journal of magnetic resonance.