Discovering High-Affinity Ligands for Proteins: SAR by NMR

Abstract

A nuclear magnetic resonance (NMR)-based method is described in which small organic molecules that bind to proximal subsites of a protein are identified, optimized, and linked together to produce high-affinity ligands. The approach is called "SAR by NMR" because structure-activity relationships (SAR) are obtained from NMR. With this technique, compounds with nanomolar affinities for the FK506 binding protein were rapidly discovered by tethering two ligands with micromolar affinities. The method reduces the amount of chemical synthesis and time required for the discovery of high-affinity ligands and appears particularly useful in target-directed drug research.

BCL-2 in the crosshairs: tipping the balance of life and death

“…This approach yielded HA14-1 (IC 50 ¼ 9 mM for competing BAK BH3/BCL-2 interaction), 103 several micromolar affinity hits from the National Cancer Institute 3D database including Compound 6, 104 A major hurdle of small molecule library screening approaches, whether virtual or biochemical, is the chemical optimization required to achieve high-potency target-binding activity from hits that are typically in the micromolar range. An alternative strategy developed by Fesik and co-workers 115 circumvents this shortcoming of library screening by chemically linking ligands that bind to adjacent sites within a target interface, effectively converting relatively low affinity interactors into conjoined high-affinity compounds. The method is called structure-activity relationships (SAR) by NMR, reflecting that compound optimization is derived from SAR determined by NMR.…”

Small-Molecule Inhibitors of IL-2/IL-2R: Lessons Learned and Applied

“…In fragment-based discovery, a compound half the size of a traditional drug (e.g., 250 Da) is screened for binding to a target of interest. Biophysical approaches are generally favored over functional assays because fragment/protein interactions tend to have low affinity, and binding methods -such as NMR (Shuker et al 1996), X-ray (Carr and Jhoti 2002), surface plasmon resonance (SPR) (Hamalainen et al 2008), or Tethering (Erlanson et al 2000) -provide additional information about binding site and/or binding stoichiometry. Active fragments are then linked or evolved in a second step and retested for binding or inhibition of the target.…”

Development of a Functionalized Xenon Biosensor

“…Steady progress is being made to incorporate NMR analysis into high-throughput and combinatorial chemistry applications. [13][14][15][16][17][18] Utilization of conventional NMR for high-throughput biosensor applications may be limited by the intrinsically low sensitivity and the complexity of spectra obtained from biomolecules and mixtures. Laser-polarization 19 of 129 Xe offers an increase in signal-tonoise by several orders of magnitude relative to the equilibrium nuclear-spin polarizations measured in normal NMR experiments.…”