Proton resonance assignments and ligand exchange kinetics in high-spin and mixed-spin myoglobin complexes using two-dimensional exchange spectroscopy.
The task of assigning resonances in proton nuclear magnetic resonance spectra of paramagnetic heme proteins can be an arduous process, but with the development of multi-dimensional NMR methods the situation has improved. It is demonstrated here that two-dimensional exchange spectroscopic experiments can be used to obtain to assignment correlations for the heme protons of methydroxy-, metthiocyano-, metaquo-, and metimidazole-myoglobin forms. All the assignments are unambiguous and straightforward when the temperature and mixing times are adjusted to minimize nuclear Overhauser cross-peaks from each complex. Moreover, saturation transfer experiments allow the study of ligand binding kinetics. The exchange rates between metaquo- and metimidazole- (or methyl substituted imidazole) myoglobin complexes are estimated. The differences between the exchange rates reflect differences in the hydrophobic and steric interactions between the ligands and the protein moiety.
Department of Chemistry, Portland State University, P.O. Box 751, Portland, OR 97207-0751, USA.
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Luo
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Peyton
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Yee
D000818:Animals; D006736:Horses; D007093:Imidazoles / Q000737:chemistry; D009211:Myoglobin / Q000737:chemistry*; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D013861:Thiocyanates / Q000737:chemistry; D014867:Water / Q000737:chemistry
D007093:Imidazoles; D009211:Myoglobin; D013861:Thiocyanates; D014867:Water; C029899:imidazole; C018100:1-methylimidazole; C023439:4-methylimidazole; C009941:potassium thiocyanate