1H NMR study of 2-methylimidazole binding to cytochrome c: a comprehensive investigation of the role of the methyl substituent on the ligand binding affinity and heme electronic structure in imidazole–cytochrome c complexes

Abstract

The binding of 2-methylimidazole (2mim) to horse heart cytochrome c (hh cyt c) has been studied by NMR spectroscopy. Some proton resonances were assigned and the kinetic and thermodynamic parameters for the binding were presented. Based on its unique hyperfine shift pattern and the anomalous temperature dependence of the heme methyl resonances, the heme electronic structure was discussed and the orientation of the bound 2mim was estimated. With these data, a comprehensive comparison of imidazole (Him), 4-methylimidazole (4mim) and 2mim bound cyt c complexes was made on the ligand binding affinity and the heme electronic structure. Different properties in these ligand–cyt c complexes provide a useful lesson for the study of protein–small molecular interactions.