The primary sequence of Cys-X-Gly-Y- (X, hydrophobic residue; Y, hydrophilic residue) is highly conserved in cytochrome P-450s. The amide NHs of Leu, Gly, and X are assumed to form NH.S hydrogen bonds which are also found in the active site fragment, Cys-Pro-Ala-Leu, of chloroperoxidase (CPO). [Fe(III)(OEP)(Z-cys-Leu-Gly-Leu-OMe)] (OEP, octaethylporphinato; Z, benzyloxycarbonyl) and [Fe(III)(OEP)(Z-cys-Pro-Ala-Leu-OMe)] were synthesized as P-450 and CPO model complexes containing the invariant amino acid fragment of the active site. The corresponding gallium(III) complexes were also synthesized to investigate the solution structures using two-dimensional (2D) NMR experiments because the Ga(III) ion is similar to the Fe(III) ion in the ionic radii and in the coordination geometry. The solution structures of the peptide part of the gallium complexes indicate that the invariant fragments maintain a beta I-turn-like conformation and then form NH.S hydrogen bonds between S(gamma)Cys and NH of the third and fourth amino acid residues. The hydrogen bonds have also been confirmed by the (2)H NMR spectra of N(2)H-substituted Fe(III) peptide complexes. The Fe(III)/Fe(II) redox potentials of the Fe(III) complexes indicate that the NH.S hydrogen bonds in the fragments causes a slight positive shift of the redox potential. The tri- and tetrapeptide Fe(III) complexes containing the invariant fragments of P-450 are kinetically stable at 30 degrees C in CH(2)Cl(2). In contrast, [Fe(III)(OEP)(Z-cys-Leu-OMe)] decomposed to give [Fe(II)(OEP)] (22%) and the corresponding disulfide immediately in CD(2)Cl(2) at 30 degrees C for 1 h. These results indicate that the invariant fragments involving the hydrogen bonds cause the stabilization of the high-spin Fe(III) resting state rather than the positive shift of Fe(III)/Fe(II) redox potential.

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