Microsecond time scale dynamics in the RXR DNA-binding domain from a combination of spin-echo and off-resonance rotating frame relaxation measurements

Slow protein dynamics can be studied by N-15 spin-echo (CPMG) and off-reson ance rotating frame relaxation through the effective field dependence of th e exchange-mediated relaxation contribution. It is shown that, by a combina tion of these complementary techniques, a more extended sampling of the mic rosecond time scale processes is achieved than by either method alone. N-15 R-2 and improved off-resonance R-1 rho experiments [Mulder et al. (1998) J . Magn. Reson., 131, 351-357] were applied to the 9-cis-retinoic acid recep tor DNA-binding domain and allowed the identification of 14 residues exhibi ting microsecond time scale dynamics. Assuming exchange between two conform ational substates, average lifetimes ranging from 37 to 416 mu s, and chemi cal shift differences of up to 3 ppm were obtained. The largest perturbatio n of tertiary structure was observed for the second zinc finger region, whi ch was found to be disordered in the solution structure [Holmbeck et al. (1 998) J. Molt Biol., 281, 271-284]. Since this zinc-coordinating domain comp rises the principal dimerization interface for RXR in a wide repertoire of complexes with different hormone receptors to their cognate response elemen ts, this finding has important implications for our understanding of nuclea r receptor assembly on DNA direct repeats. The flexibility observed for the dimerization domain may explain how RXR, through the ability to adaptively interact with a wide variety of highly homologous partner molecules, demon strates such a versatile DNA-binding repertoire.