NMR spectra and protein-protein interactions. (a) A one-dimensional NMR spectrum of a globin from a marine bloodworm. The spectrum represents the amount of chemical shift for each proton in a peptide segment. For a protein, the proton signals do not resolve in a one-dimensional spectrum, as indicated by the many overlapping peaks. (b) A two-dimensional NMR spectrum of the same globin molecule. The spots and their intensities that lie along the diagonal line are equivalent to the data contained in the peaks of the one-dimensional spectrum. The off-diagonal peaks (e.g., peaks 1 and 2) are nuclear Overhauser effect (NOE) signals generated by close-range interactions of ¹H atoms that generate signals quite distant in the one-dimensional spectrum. (c) The two-dimensional COSY analysis identifies proton-proton signals through one or two covalent bonds (“through-bond” signals) and thus is limited to individual amino acid units. (d) The NOESY analysis yields NOE signals resulting from proton-proton interactions occurring through empty space (“through-space” signals) and thus identifies protons close in space but not necessarily close in the primary sequence.