Characterization by NMR of the heme-myoglobin adduct formed during the reductive metabolism of BrCCl3. Covalent bonding of the proximal histidine to the ring I vinyl group

• Published in: The Journal of biological chemistry Vol. 266; no. 5; pp. 3208 - 3214
• Main Authors: Osawa, Y, Highet, R J, Bax, A, Pohl, L R
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 15-02-1991
• Subjects: Heme - metabolism; Histidine - chemistry; Hydrocarbons, Brominated - metabolism; Magnetic Resonance Spectroscopy; Models, Molecular; Myoglobin - metabolism; Oxidation-Reduction; Spectrophotometry, Ultraviolet; Trihalomethanes
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)49975-9

The reductive debromination of BrCCl3 by ferrous deoxymyoglobin leads to the covalent bonding of the prosthetic heme to the
protein. We have previously shown, by the use of peptide mapping and mass spectrometry, that histidine residue 93 is covalently
bound to the heme moiety. In the present study the structure of the heme adduct was more completely determined by 1H and 13C
NMR techniques. We have found that the ring I vinyl group of the prosthetic heme was altered by the addition of a histidine
imidazole nitrogen to the alpha-carbon and a CCl2 moiety to the beta-carbon. The electronic absorption spectra of the oxidized
and reduced states of the altered heme-protein indicated that the heme-iron exists in a bis-histidine-ligated form. Analysis
of the crystal structure of native myoglobin suggested that for the altered heme-protein, histidine residues 97 and 64 are
ligated to the heme-iron and that residue 97 has replaced the native proximal histidine residue 93. These movements, in effect
a "histidine shuffle" at the active site, may be responsible for the enhanced reducing activity of the altered protein.