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Abstract

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.

The Src Homology 3 Domain Is Required for Junctional Adhesion Molecule Binding to the Third PDZ Domain of the Scaffolding Protein ZO-1

“…Experimental conditions were 10 M protein in 100 mM PO 4 , pH 7.2, in 90% 1 H 2 O, 10% 2 H 2 O at 25°C in 3-mm NMR tubes. The WaterLOGSY experiments were performed as described previously (29). The data were collected with a sweep width of 14,370 Hz and a relaxation delay of 2 s. Water was selectively saturated using a 2-ms square-shaped pulse with a mixing time of 2 s. All data were processed by NMRPipe (30) with referencing to the water peak at 4.773 ppm and using a 4-Hz line-broadening window.…”

“…S4). To probe for affinity differences between JAM-A_P12 and JAM-A_P6, we performed a WaterLOGSY-based titration (29). We determined the dissociation constant (K d ) for the peptide from PSG.…”

Discovery of novel FabF ligands inspired by platensimycin by integrating structure-based design with diversity-oriented synthetic accessibility

“…The amide 7 was prepared (Scheme 5) as a reference ligand for a WaterLOGSY assay [50][51][52] to evaluate the de novo designed ligands.…”

Application of residual dipolar couplings in organic compounds

“…This fast exchange allows ligand-based high-resolution techniques, 91 such as transferred NOE, 92 saturation transfer difference (STD) 93,94 and water-ligand observed via gradient spectroscopy experiments (WaterLOGSY), 95,96 to study ligand-receptor interactions and structure-activity relationships (SAR). But without the structure, or at least the NH assignments of the protein itself, the relative geometry of bound ligand against protein is difficult to determine from these techniques.…”