Heat capacities and a snapshot of the energy landscape in protein GB1 from the pre-denaturation temperature dependence of backbone NH nanosecond fluctuations.
Protein stability is usually characterized calorimetrically by a melting temperature and related thermodynamic parameters. Despite its importance, the microscopic origin of the melting transition and the relationship between thermodynamic stability and dynamics remains a mystery. Here, NMR relaxation parameters were acquired for backbone 15NH groups of the 56 residue immunoglobulin-binding domain of streptococcal protein G over a pre-denaturation temperature range of 5-50 degrees C. Relaxation data were analyzed using three methods: the standard three-Lorentzian model free approach; the F(omega)=2omegaJ(omega) spectral density approach that yields motional correlation time distributions, and a new approach that determines frequency-dependent order parameters. Regardless of the method of analysis, the temperature dependence of internal motional correlation times and order parameters is essentially the same. Nanosecond time-scale internal motions are found for all NHs in the protein, and their temperature dependence yields activation energies ranging up to about 33kJ/mol residue. NH motional barrier heights are structurally correlated, with the largest energy barriers being found for residues in the most "rigid" segments of the fold: beta-strands 1 and 4 and the alpha-helix. Trends in this landscape also parallel the free energy of folding-unfolding derived from hydrogen-deuterium (H-D) exchange measurements, indicating that the energetics for internal motions occurring on the nanosecond time-scale mirror those occurring on the much slower time-scale of H-D exchange. Residual heat capacities, derived from the temperature dependence of order parameters, range from near zero to near 100J/mol K residue and correlate with this energy landscape. These results provide a unique picture of this protein's energy landscape and a relationship between thermodynamic stability and dynamics that suggests thermosensitive regions in the fold that could initiate the melting process.
Idiyatullin
Djaudat
D
Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, 6-155 Jackson Hall, 321 Church Street, Minneapolis, MN 55455, USA.
Nesmelova
Irina
I
Daragan
Vladimir A
VA
Mayo
Kevin H
KH
D001426:Bacterial Proteins / Q000737:chemistry*; D004734:Energy Metabolism; D019277:Entropy; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D011489:Protein Denaturation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D013696:Temperature; D013816:Thermodynamics; D013997:Time Factors
D001426:Bacterial Proteins; C034284:IgG Fc-binding protein, Streptococcus