Determination of the Three-Dimensional Structure of a New Crystalline Form of N-Acetyl-Pro-Gly-Phe As Revealed by 13C REDOR, X-Ray Diffraction, and Molecular Dynamics Calculation

The interatomic distances in the crystalline specimen of 13 C, 15 N doubly labeled peptides [1- 13 C] N -acetyl-Pro-[ 15 N]Gly-Phe ( I ), N -acetyl-[1- 13 C]Pro-Gly-[ 15 N]Phe ( II ), and [1- 13 C] N -acetyl-Pro-Gly-[ 15 N]Phe ( III ) evaluated from rotational echo double resonance (REDOR) data were compared with those from X-ray diffraction studies and justify our novel approach. The minimization of B 1 inhomogeneity was critical to obtain accurate distances, which were achieved by confinement of the samples in the central portion (50% of the total filling volume of the rotor). The effect of the finite length of the π pulse was found to be negligible as long as the pulse length is less than 10% of the rotor cycle. The 13 C··· 15 N distances obtained from 13 C REDOR were thus 3.24 ± 0.05, 3.43 ± 0.05, and 4.07 ± 0.05 Å for I, II, and III, respectively. The REDOR-derived conformation of this peptide was β-turn type I, consistent with our X-ray diffraction study (orthorhombic crystal). The maximum deviation of the distances determined by NMR and X-ray diffraction is 0.08 Å despite the complete neglect of the dipolar interactions with the labeled nuclei of neighboring molecules and natural abundance nuclei. The precision and accuracy given by 13 C REDOR experiments are on the order of ±0.05 Å. Distinction between the two types of β-turn forms including the β-turn type II found in the monoclinic crystal of this peptide whose interatomic distances are different by about 0.57 Å is made possible only by the very accurate REDOR measurement. Finally, we found that the three-dimensional structure of this peptide was well reproduced by a molecular dynamics simulation by taking into account all the intermolecular interactions in the crystals.