hnca nmr assignment

Protein NMR

A practical guide.

Reference: R.T. Clubb, V. Thanabal and G. Wagner (1992) J. Magn. Reson. 97 213-217. ( Link to Article )

Minimum labelling: 15 N, 13 C

Dimensions: 3

The Magnetisation is transferred from 1 H to 15 N and then via the N-Cα J-coupling to the 13 Cα. From there it is transferred to the 13 CO via the 13 Cα- 13 CO J-coupling. For detection the magnetisation is transferred back the same way: from 13 CO to 13 Cα, 15 N and finally 1 H. The chemical shift is only evolved on 1 H, 15 N and 13 CO and not on the 13 Cα. The result is a three-dimensional spectrum. Because the amide nitrogen is coupled both to the Cα of its own residue and that of the preceding residue, both these transfers occur and transfer to both 13 CO nuclei occurs. Thus for each NH group, two carbonyl groups are observed in the spectrum. But because the coupling between N i and Cα i is stronger than that between N i and Cα i-1 , the H i -N i -CO i peak generally ends up being more intense than the H i -N i -CO i-1 peak.

This experiment can be useful for backbone assignment when used in conjunction with the HNCA, HN(CO)CA and HNCO if the CBCANNH and CBCA(CO)NNH spectra are of bad quality.

An overlay of the HNCO and HN(CA)CO spectra makes it very easy to distinguish between CO i and CO i-1 for each NH group.