Methods for sequential resonance assignment in solid, uniformly 13C, 15N labelled peptides:: Quantification and application to antamanide

The application of adiabatic polarization-transfer experiments to resonance assignment in solid, uniformly C-13-N-15-labelled polypeptides is demonstrated for the cyclic decapeptide antamanide. A homonuclear correlation experiment employing the DREAM sequence for adiabatic dipolar transfer yields a complete assignment of the C-alpha and aliphatic side-chain C-13 resonances to amino acid types. The same information can be obtained from a TOBSY experiment using the recently introduced P9(12)(1) TOBSY sequence, which employs the J couplings as a transfer mechanism. A comparison of the two methods is presented. Except for some aromatic phenylalanine resonances, a complete sequence-specific assignment of the C-13 and N-15 resonances in antamanide is achieved by a series of selective or broadband adiabatic triple-resonance experiments. Heteronuclear transfer by adiabatic-passage Hartmann-Hahn cross polarization is combined with adiabatic homonuclear transfer by the DREAM and rotational-resonance tickling sequences into two- and three-dimensional experiments. The performance of these experiments is evaluated quantitatively.