Off-resonance TROSY (R_1ρ - R₁) for quantitation of fast exchange processes in large proteins

Current solution NMR experiments for characterizing conformational exchange processes in large proteins are limited to exchange rates ca. 500-3000 s^-1. A TROSY-based constant relaxation time (R_1ρ - R₁) experiment is designed to extend this capability to measure motion with rates up to 10⁵ s^-1 in large macromolecules. The experiment combines off-resonance spin-lock rf fields, which provide access to the faster time-scale dynamics, with TROSY coherence selection, which extends the molecular-weight range available for study. When implemented on the 53-kDa dimeric enzyme triosephosphate isomerase, the experiment yielded substantial gains in signal-to-noise (up to 60%) over current experiments at modest static magnetic fields (14.1 T). The TROSY (R_1ρ - R₁) experiment should therefore be of general utility for investigation of fast conformational exchange events in large proteins.