title: Thermal unfolding of ribonuclease T1 studied by multi-dimensional NMR spectroscopy.
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.
authors.affiliation: School of Life Science, Tokyo University of Pharmacy and Life Science, Hachioji, Tokyo 192-0392, Japan.
authors.forename: Hisae
authors.initials: H
authors.lastname: Matsuura
authors.forename: Sakurako
authors.initials: S
authors.lastname: Shimotakahara
authors.forename: Chiseko
authors.initials: C
authors.lastname: Sakuma
authors.forename: Mitsuru
authors.initials: M
authors.lastname: Tashiro
authors.forename: Heisaburo
authors.initials: H
authors.lastname: Shindo
authors.forename: Kiwa
authors.initials: K
authors.lastname: Mochizuki
authors.forename: Akihiko
authors.initials: A
authors.lastname: Yamagishi
authors.forename: Masaki
authors.initials: M
authors.lastname: Kojima
authors.forename: Kenji
authors.initials: K
authors.lastname: Takahashi
mesh_terms: D000465:Algorithms; D004926:Escherichia coli / Q000737:chemistry / Q000201:enzymology; D006358:Hot Temperature; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D011487:Protein Conformation; D017510:Protein Folding; D006163:Ribonuclease T1 / Q000737:chemistry*; D013696:Temperature; D013816:Thermodynamics
chemical_list: D006163:Ribonuclease T1