Improved measurement of ¹⁵ N-{ ¹ H} NOEs in the presence of H(N)-water proton chemical exchange

A simple method is presented to accurately determine ¹⁵ N-{ ¹ H} NOEs in biomolecules in the presence of H(N)-water proton chemical exchange. Three measurements are required: one with nonselective proton saturation and two with different water saturation conditions to determine the equilibrium value of the ¹⁵ N signal. This approach is exemplified with data on two peptides, one helix-forming 17-mer and one compactly folded 56-mer. Results indicate that ¹⁵ N-{ ¹ H} NOEs determined using the standard approach with short recycle times (3 to 4 s) can be significantly in error when H(N)-water proton chemical exchange is relatively rapid, water proton relaxation is relatively slow, and ¹⁵ N-{ ¹ H} NOEs are away from the value of -1. This new method avoids such inaccuracies resulting from the use of short recycle times.