COMPARISON OF N-15-DETERMINED AND C-13-DETERMINED PARAMETERS OF MOBILITY IN MELITTIN

Backbone and tryptophan side-chain mobilities in the 26-residue, cytol ytic peptide melittin (MLT) were investigated by N-15 and C-13 NMR. Sp ecifically, inverse-detected N-15 T-1 and steady-state NOE measurement s were made at 30 and 51 MHz on MLT at 22 degrees C enriched with N-15 at six amide positions and in the Trp(19) side chain. Both the disord ered MLT monomer (1.2 mM peptide at pH 3.6 in neat water) and alpha-he lical MLT tetramer (4.0 mM peptide at pH 5.2 in 150 mM phosphate buffe r) were examined. The relaxation data were analyzed in terms of the Li pari and Szabo model-free formalism with three parameters: tau(m), the correlation time for the overall rotation; S-2, a site-specific order parameter which is a measure of the amplitude of the internal motion; and tau(e), a local, effective correlation time of the internal motio n. A comparison was made of motional parameters from the 15N measureme nts and from C-13 measurements on MLT, the latter having been made her e and previously [Kemple et al. (1997) Biochemistry, 36, 1678-1688]. t au(m) and tau(e) values were consistent from data on the two nuclei. I n the MLT monomer, S-2 values for the backbone N-H and C alpha-H vecto rs in the same residue were similar in value but in the tetramer the N -H order parameters were about 0.2 units larger than the C alpha-H ord er parameters. The Trp side-chain N-H and C-H order parameters, and ta u(e) values were generally similar in both the monomer and tetramer. I mplications of these results regarding the dynamics of MLT are examine d.