Ligand-binding and electrochemical properties of 5-coordinate imidazole-ligated Fe(II) porphyrins are of intense contemporary interest in areas from biomimetic chemistry to chemical sensors, non-cryogenic air separation, and Pt-free catalysts for electrochemical reduction of oxygen. The 5-coordinate state, however, is not easily achieved in synthetic compounds because binding of a 2^nd imidazole to a ferroheme proceeds with a higher affinity than binding of the first: i.e., mono-imidazole ligated ferroheme is metastable with respect to a mixture of the coordinatively saturated 6-coordinate and catalytically inactive 4-coordinate hemes. The most certain strategy to overcome this intrinsic difference in affinities has been to tether an imidazole to the porphyrin to achieve intramolecular ligation enhanced by the chelate effect, but such complexes are hard to synthesize and are unsuitable for practical applications. Simple structural motifs in which the mono-imidazole ligation of ferroheme can be enforced are needed. In addition, cooperative O₂ binding is a property that is particularly attractive for non-cryogenic separation of O₂ from air, as it decreases the pressure difference at which one achieves useful degree of oxygenation and deoxygenation of the O₂ carrier as illustrated by hemoglobin. Prior to our work cooperative O₂ binding to  synthetic hemes was only achieved in crystals, in polymers, or in the presence of a large excess of competing ligands, i.e., imidazole.

We demonstrated a simple structural motif that leads to stable 5-coordinate mono-imidazole ligated Fe(II) centers in solution in the absence of exogenous ligands and carried out experimental and computational studies of these systems. Spontaneous dimerization of two imidazolyl-containing Fe(II) porphyrins 1Fe or 2Fe resulted in a pair of 5-coordinate imidazole-ligated Fe(II) centers. A variety of small ligands (imidazole, pyridine, nitrosoaryls and isocyanides) bind to these centers with high affinity and without causing dimer dissociation. Moreover, the binding is cooperative: for example, (1Fe)₂ binds a 2^nd isocyanide with ~50-fold higher affinity than the first, corresponding to the allosteric free energy of interaction (DG_a) of 13±3 kJ/mol, which is comparable to that of hemoglobin!

We synthesized monomeric bases 1H₂-5H₂ in one- to three-step procedures and metallated them with Fe(II), Mg(II) and Zn(II). All compounds were characterized by UV-vis and NMR spectroscopies. Stable dimers were observed for Fe, Zn and Mg complexes of porphyrin 1-3 (dimerization constants up to 5´10⁸ M^-1) in non-coordinating solvents, whereas only Zn and Mg complexes of 4 and 5 formed dimers with low stability.

Ligand binding to (1Fe)₂ and (2Fe)₂ was cooperative, with DG_a varying from 13±3 kJ/mol for isocyanide binding to 5±2 kJ/mol for imidazole (corresponding Hill coefficients from 2 to 1.5) The binding constants and allosteric interaction energies were independent of the solvent and the dimer concentration. Our dimers (1Fe)₂ and (2Fe)₂ present the simplest known hemes that bind ligands cooperatively. Remarkably, the two Fe(II) centers in (3Fe)₂ bound the same ligands completely independently!

Relatively small size of our complexes allowed us to study dimers (1Fe)₂ and (3Fe)₂ and their adducts with MeNC by electronic structure computations at the B3LYP/6-31g level to understand the structural and electronic changes that occur upon ligand binding. Our benchmarking studies for monomeric porphyrins in relevant ligation states proved that the selected model chemistry was adequate for describing the structural parameters and electronic state of (1Fe)₂ and (3Fe)₂ and their adducts. We established procedures to overcome difficulties related to the high-spin state of 5-coordinate Fe(II) centers in the parent dimers and dimers with a single exogenous ligand. Computations showed limited structural and electronic communication between the two Fe cites in either dimer. The electronic and structural changes at the binding site are pronounced: it converts from the high-spin to the low-spin state, the Fe atom moves into the porphyrin plane by up to 0.35 Å and the Fe-N_imidazole bond shortens by up to 0.06 Å. The spectator site however remains essentially unaffected both in the first and the second binding event. These results indicate a large contribution of the entropic term to the site-site interaction energy in (1Fe)₂. These DFT computations suggest that the principal difference between the dimers with phenyl-imidazole links (1Fe)₂ and with imidazole links (3Fe)₂ is destabilization of (3Fe)₂ in the dimeric form by the steric repulsion between the two porphyrin planes so close together (calculated interplane separation for model (3Fe)₂ was 3.2-3.5 Å depending on the ligation state).

In addition to the two publications already accepted or in press, work partially supported by this grant resulted in a paper currently under review at J. Phys. Chem. A (“DFT calculations of the lowest-energy quintet and triplet states of model hemes: effects of functional, basis set and zero-point energy corrections” by D. Khvostichenko, A. Choi and R. Boulatov) and a paper in submission to Chem. Comm. (“Simple dimers containing dissociatively-stable mono-imidazole ligated ferrohemes” by Q. Yang, D. Khvostichenko, J. Atkinson and R. Boulatov; porphyrins 3Fe-5Fe).