Distal and Proximal Ligand Interactions in Heme Proteins: Correlations between C–O and Fe–C Vibrational Frequencies, Oxygen-17 and Carbon-13 Nuclear Magnetic Resonance Chemical Shifts, and Oxygen-17 Nuclear Quadrupole Coupling Constants in C¹⁷O- and ¹³CO-Labeled Species

We have obtained the oxygen-17 nuclear magnetic resonance (NMR) spectra of a variety of C^l7O-labeled heme proteins, including sperm whale (Physeter catodon) myoglobin, two synthetic sperm whale myoglobin mutants (His E7 → Val E7; His E7 → Phe E7), adult human hemoglobin, rabbit (Oryctolagus cuniculus) hemoglobin, horseradish (Cochlearia armoracia) peroxidase (E.C. 1.11.1.7) isoenzymes A and C, and Caldariomyces fumago chloroperoxidase (E.C. 1.11.1.10), in some cases as a function of pH, and have determined their isotropic ¹⁷O NMR chemical shifts, δ_i, and spin-lattice relaxation times, T₁. We have also obtained similar results on a picket fence prophyrin, [5, 10, 15, 20-tetrakis(α,α,α,α-pivalamidophenyl)porphyrinato]iron(II) (l-Melm)CO, both in solution and in the solid state. Our results show an excellent correlation between the infrared C–O vibrational frequencies, v(C–O), and δ_i, between μ(C–O) and the ^,7O nuclear quadrupole coupling constant (e²qQ/h, derived from T₁), and as expected between e²qQ/h and δ_i. Taken together with the work of others on the ¹³C NMR of ¹³CO-labeled proteins, where we find an excellent correlation between δ_i(¹³C) and v(Fe–C), our results suggest that IR and NMR measurements reflect the same interaction, which is thought to be primarily the degree of π-back-bonding from Fe d to CO π* orbitals, as outlined previously [Li, X.-Y., & Spiro, T. G. (1988) J. Am. Chem. Soc. 110, 6024], The modulation of this interaction by the local charge field of the distal heme residue (histidine, glutamine, arginine, and possibly lysine) in a variety of species and mutants, as reflected in the NMR and IR measurements, is discussed, as is the effect of cysteine as the proximal heme ligand.