An experimental and quantum chemical investigation of CO binding to heme proteins and model systems: A unified model based on ¹³C, ¹⁷O, and ⁵⁷Fe nuclear magnetic resonance and⁵⁷Fe mossbauer 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), ⁵⁷Fe Mossbauer, and infrared spectroscopic techniques, combined with density functional theoretical calculations to analyze the spectroscopic results. The results of ¹³C NMR isotropic chemical shift, ¹³C NMR chemical shift anisotropy, ¹⁷O NMR isotropic chemical shift, ¹⁷O nuclear quadrupole coupling constant, ⁵⁷Fe NMR isotropic chemical shift, ⁵⁷Fe Mossbauer quadrupolar splitting, and infrared measurements indicate that CO binds to Fe in a close to linear fashion in all conformational substates. The ¹³C-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 ¹⁷O chemical shift, and the ¹⁷O 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 V(co) for the model compound of 1969 cm^-1 is also that found for A(o) proteins. Results for the A₁ substate (including the ⁵⁷Fe NMR chemical shift and Mossbauer 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 ¹³C shift, ¹³C shift anisotropy, ¹⁷O shift, ¹⁷O NQCC, ⁵⁷Fe shift, ⁵⁷Fe Mossbauer quadrupole splitting, and v(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.