Unfolding of the loggerhead sea turtle (Caretta caretta) myoglobin: A (1)H-NMR and electronic absorbance study.
The effect of urea concentration on the backbone solution structure of the cyanide derivative of ferric Caretta caretta myoglobin (at pH 5.4) is reported. By addition of urea, sequential and long-range nuclear Overhauser effects (NOEs) are gradually lost. By using the residual NOE constraints to build the molecular model, a picture of the unfolding pathway was obtained. When the urea concentration is raised to 2.2 M, helices A and B appear largely disordered; helices C, D, and F loose structural constraints at 3.0 M urea. At urea concentration >6 M, the protein appears to be fully unfolded, including the GH hairpin and helix E stabilizing the prosthetic group. Reversible and cooperative denaturation isotherms obtained by following NOE peaks are considerably different from those obtained by monitoring electronic absorption changes. The reversible and cooperative urea-dependent folding-unfolding process of C. caretta myoglobin follows the minimum three-state mechanism N long left and right arrow X long left and right arrow D, where X represents a disordered globin structure (occurring at approximately 4 M urea) that still binds the heme.
Department of Chemistry "IFM," University of Torino, I-10125 Torino, Italy.
Daniela Delli
DD
Castelli
Elena
E
Lovera
Paolo
P
Ascenzi
Mauro
M
Fasano
D000818:Animals; D009211:Myoglobin / Q000737:chemistry*; D019906:Nuclear Magnetic Resonance, Biomolecular; D011489:Protein Denaturation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D013816:Thermodynamics; D014426:Turtles; D014508:Urea / Q000737:chemistry
D009211:Myoglobin; D014508:Urea