Measurement of slow (mu s-ms) time scale dynamics in protein side chains by N-15 relaxation dispersion NMR spectroscopy: Application to Asn and Gln residues in a cavity mutant of T4 lysozyme

A new NMR experiment is presented for the measurement of mus-ms time scale dynamics of Asn and Gin side chains in proteins. Exchange contributions to the N-15 line widths of side chain residues are determined via a relaxation dispersion experiment in which the effective nitrogen transverse relaxatio n rate is measured as a function of the number of refocusing pulses in cons tant-time, variable spacing CPMG intervals. The evolution of magnetization from scalar couplings and dipole-dipole cross-correlations, which has limit ed studies of exchange in multi-spin systems in the past, does not affect t he extraction of accurate exchange parameters from relaxation profiles of N H2 groups obtained in the present experiment. The utility of the method is demonstrated with an application to a Leu --> Ala cavity mutant of T4 lysoz yme, L99A. II is shown that many of the side chain amide groups of Asn and Gin residues in the C-terminal domain of the protein are affected by a chem ical exchange process which may be important in facilitating the rapid bind ing of hydrophobic ligands to the cavity.