Proton‐detected C,H correlation NMR techniques for the complete assignment of all proton and carbon resonances of a cyclic peptide

The assignment of the proton and carbon spectra of the cyclic peptide cyclo(‐Phe¹‐Pro²‐Thr³‐Lys(Z)⁴‐Trp⁵‐Phe⁶‐) was accomplished by the application of multiple quantum proton‐detected heteronuclear correlation spectroscopy. Since the proton spectrum shows severe overlap, the carbon chemical shifts were used to disentangle the proton resonances. The methodology described is useful even in cases where only limited quantities of materials are available. The combination of (i) a proton decoupled C,H correlation spectrum and (ii) a (not decoupled) H‐relayed C,H correlation gave the assignments of all CH_n groups. The non‐protonated carbons, i.e. the carbonyl carbons of the peptide bond, were assigned with a C,H correlation optimized for long‐range couplings, an experiment that also gave helpful information about the conformational features of the hexapeptide. The cyclic peptide contains a Phe‐Pro cis‐peptide bond forming a βVI‐like bend and a β‐turn about the amino acids Thr‐Lys‐Trp‐Phe. Although the conventional discussion of NMR parameters indicates a strong preference for one conformation, the quantitative evaluation of NOE‐derived distances in restrained MD calculations proves that the type of β‐turn in the last‐mentioned region is not unique. Whereas the MD calculations converge to a βII' turn, the vicinal proton coupling constants are in better agreement with type βI. Thus a dynamic equilibrium of the backbone is proposed.