An Experimental and Quantum Chemical Investigation of CO Binding to Heme Proteins and Model Systems:  A Unified Model Based on¹³C,¹⁷O, and⁵⁷Fe Nuclear Magnetic Resonance and⁵⁷Fe Mössbauer and Infrared Spectroscopies

We have investigated the question of how CO ligands bind to iron in metalloporphyrins and metalloproteins by using a combination of nuclear magnetic resonance (NMR), 57 Fe Mössbauer, and infrared spectroscopic techniques, combined with density functional theoretical calculations to analyze the spectroscopic results. The results of 13 C NMR isotropic chemical shift, 13 C NMR chemical shift anisotropy, 17 O NMR isotropic chemical shift, 17 O nuclear quadrupole coupling constant, 57 Fe NMR isotropic chemical shift, 57 Fe Mössbauer quadrupolar splitting, and infrared measurements indicate that CO binds to Fe in a close to linear fashion in all conformational substates. The 13 C-isotropic shift and shift anisotropy for an A o substate model compound: Fe(5,10,15,20-tetraphenylporphyrin)(CO)( N -methylimidazole), as well as the 17 O chemical shift, and the 17 O nuclear quadrupole coupling constant (NQCC) are virtually the same as those found in the A o substate of Physeter catodon CO myoglobin and lead to most probable ligand tilt (τ) and bend (β) angles of 0° and 1° when using a Bayesian probability or Z surface method for structure determination. The infrared ν CO for the model compound of 1969 cm - 1 is also that found for A o proteins. Results for the A 1 substate (including the 57 Fe NMR chemical shift and Mössbauer quadrupole splitting) are also consistent with close to linear and untilted Fe−C−O geometries (τ = 4°, β = 7°), with the small changes in ligand spectroscopic parameters being attributed to electrostatic field effects. When taken together, the 13 C shift, 13 C shift anisotropy, 17 O shift, 17 O NQCC, 57 Fe shift, 57 Fe Mössbauer quadrupole splitting, and ν CO all strongly indicate very close to linear and untilted Fe−C−O geometries for all carbonmonoxyheme proteins. These results represent the first detailed quantum chemical analysis of metal−ligand geometries in metalloproteins using up to seven different spectroscopic observables from three types of spectroscopy and suggest a generalized approach to structure determination.