2D-NMR studies of the effects of axial substitution on two helices in horse cytochrome c.

The sequential amino acids from Gly-1 to Cys-14 in the N-terminal segment and from Lys-88 to Glu-104 in the C-terminal segment of cytochrome c-imidazole complex (Im-cyt c) have been studied by two-dimensional nuclear magnetic resonance techniques. Resonance assignments for the main-chain and side-chain protons are reported. Qualitative interpretation of nuclear Overhauser enhancement data allows the secondary structure of the two segments to be described, which indicate the patterns of NOEs found are consistent with an alpha-helix between Val-3 and Cys-14 in the N-terminus, and between Lys-88 and Asn-103 in the C-terminus. Two alpha-helices are found to be maintained. Comparison of the long-range NOEs of Im-cyt c relative to the crystal structure of native cytochrome c reveals apparent conformational changes of some side-chains especially those close to heme pocket within the N- and C-terminal helices resulting from the binding of imidazole to iron by displacing native Met-80 side-chain. The explanation for ligand-induced changes within the N- and C-helices is therefore suggested.