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Updated: Aug 20, 2026

NMR 15N Relaxation Experiments for the Investigation of Picosecond to Nanoseconds Structural Dynamics of Proteins
Published on: November 1, 2024
Thermal unfolding of ribonuclease T1 studied by multi-dimensional NMR spectroscopy
Hisae Matsuura1, Sakurako Shimotakahara, Chiseko Sakuma
1School of Life Science, Tokyo University of Pharmacy and Life Science, Hachioji, Tokyo 192-0392, Japan.
Abstract:
Thermal unfolding of ribonculease (RNase) T1 was studied by 1H nuclear Overhauser enhancement spectroscopy (NOESY) and 1H- 15N heteronuclear single-quantum coherence (HSQC) NMR spectroscopy at various temperatures. Native RNase T1 is a single-chain molecule of 104 amino acid residues, and has a single alpha-helix and two beta-sheets, A and B, which consist of two and five strands, respectively. Singular value decomposition analysis based on temperature-dependent HSQC spectra revealed that the thermal unfolding of RNase T1 can be described by a two-state transition model. The midpoint temperature and the change in enthalpy were determined as 54.0 degrees C and 696 kJ/mol, respectively, which are consistent with results obtained by other methods. To analyze the transition profile in more detail, we investigated local structural changes using temperature-dependent NOE intensities. The results indicate that the helical region starts to unfold at lower temperature than some beta-strands (B3, B4, and B5 in beta-sheet B). These beta-strands correspond to the hydrophobic cluster region, which had been expected to be a folding core. This was confirmed by structure calculations using the residual NOEs observed at 56 degrees C. Thus, the two-state transition of RNase T1 appears to involve locally different conformational changes.
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