Long-distance rotational echo double resonance measurements for the determination of secondary structure and conformational heterogeneity in peptides

Abstract

The utility of rotational echo double resonance (REDOR) NMR spectroscopy for determining the conformations of linear peptides has been examined critically using a series of crystalline and amorphous samples. The focus of the present work was the evaluation of long-distance (>5 Å) interactions using 13 C– 15 N dephasing. Detailed studies of specifically labeled melanostatin and synthetic analogs of the α-factor yeast mating hormone show that nitrogen-dephased, carbon-observe REDOR measurements are reliable for distances up to 6.0 Å, and that dipolar interactions can be detected for distances up to 7 Å. By contrast, nitrogen-observe REDOR gives reliable results only for distances shorter than 5.0 Å. To measure distances accurately, REDOR data must be corrected for the effects of natural-abundance spins. These corrections are particularly important for measuring long distances, which are of the greatest value for determining peptide secondary structure. We have developed a spherical shell model for calculating the effect of these background spins. The REDOR studies also indicate that in a lyophilized powder, the tridecapeptide α-factor mating pheromone from Saccharomyces cerevisiae (WHWLQLKPGQPMY) probably exists as a distribution of different turn structures around the KPGQ region. This finding revises previous solid-state NMR studies on this peptide, which concluded α-factor assumes a distorted type-I β-turn in the Pro-Gly central region of the molecule [J.R. Garbow, M. Breslav, O. Antohi, F. Naider, Biochemistry, 33 (1994) 10094].