In this chapter, typical peptide‐specific properties are highlighted, including lack of secondary and tertiary structure, high solvent exposure, small core volume, natural isotope abundance, and low spectral crowding. NMR experiments and structure calculation protocols developed for the investigation of peptides with distinct preferred conformations, as observed in constrained (cyclic or N‐methylated) peptides, are also presented. Many of these approaches differ greatly from NMR experiments and structure calculation strategies designed for studying proteins, as demonstrated by the following:

  1. Heteronuclear multiple‐bond correlations (HMBCs) play an important role in the assignment process and in determining preferred dihedral angles.
  2. Rotating frame Overhauser effect spectroscopy (ROESY) rather than nuclear Overhauser effect spectroscopy (NOESY) spectra are used for the calculation of distance restraints that are obtained with a higher precision than distance restraints in proteins.
  3. math coupling constants can often be read from different kinds of multiplet splittings (e.g., from one‐dimensional math NMR spectra or from correlation spectroscopy (COSY)‐type multiplet patterns).

Different types of ROESY experiments, methods for the determination of math coupling constants, as well as an overview of residual dipolar coupling‐based peptide NMR studies and the alignment media used therein are described. The impact of peptide flexibility for conformational studies is discussed in the context of distance and dihedral restraints. Furthermore, a common peptide structure calculation protocol including distance geometry and molecular dynamics calculations is presented.