Measuring ligand-protein binding using NMR diffusion experiments

Characterization of ligand‐protein interactions is important because many biologically important processes are mediated by the binding of a small molecule to an enzyme or cell‐surface receptor. Pulsed‐field gradient nuclear magnetic resonance (PFG‐NMR) spectroscopy measurements of diffusion coefficients permit noninvasive, quantitative analysis of binding over a broad range of dissociation constants and ligand:protein concentration ratios. An arsenal of specific and selective PFG‐NMR methods has been developed by exploiting differential behaviors of small molecule ligands and macromolecular proteins. A discussion of several PFG‐NMR methods and their relevant applications reveals the analytical value of using PFG‐NMR diffusion measurements for studying ligand‐protein binding. © 2004 Wiley Periodicals, Inc. Concepts Magn Reson Part A 20A: 24–41, 2004.