A new amide proton R1ρ experiment permits accurate characterization of microsecond time-scale conformational exchange

A new off-resonance spin-lock experiment to record relaxation dispersion profiles of amide protons is presented. The sensitivity-enhanced HSQC-type sequence is designed to minimize the interference from cross-relaxation effects and ensure that the dispersion profiles in the absence of mu s-ms time-scale dynamics are. at. Toward this end (i) the proton background is eliminated by sample deuteration (Ishima et al., 1998), (ii) H-1 spin lock is applied to two-spin modes 2(H-x sin theta + H-z cos theta)N-z, and (iii) the tilt angle theta approximate to 35 degrees is maintained throughout the series of measurements (Desvaux et al., 1995). The relaxation dispersion profiles recorded in this manner sample a wide range of effective rf field strengths ( up to and in excess of 20 kHz) which makes them particularly suitable for studies of motions on the time scale <= 100 mu s. The new experiment has been tested on the Ca2+- loaded regulatory domain of cardiac troponin C. Many residues show pronounced dispersions with remarkably similar correlation times of similar to 30 mu s. Furthermore, these residues are localized in the regions that have been previously implicated in conformational changes (Spyracopoulos et al., 1997).