Sequential 1H‐NMR assignment and solution structure of bovine pancreatic ribonuclease A

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Abstract
Assignments for 1H‐NMR resonances of most of the residues of bovine pancreatic ribonuclease (RNase A) have been obtained by sequence‐specific methods. Identification and classification of spin systems have been carried out by two‐dimensional phase‐sensitive correlated spectroscopy (360 MHz) and single relayed coherence transfer spectroscopy. Sequence‐specific assignments have been achieved by phase‐sensitive two‐dimensional nuclear Overhauser effect spectroscopy. To overcome the problem of spectral overlap use has been made of (a) an exhaustive analysis of partly exchanged RNase A (spectra in D2O), (b) a comparison with the subtilisin‐modified enzyme (RNase S) and (c) small spectral perturbations caused by changes in pH and temperature. The secondary structure elements have been identified from the observed sequential, medium and long‐range nuclear Overhauser effects together with data from amide‐exchange rates. All information collected leads to the conclusion that the crystal and the solution structures are closely similar.