Kinetics and mechanism studies in biomimetic chemistry: metalloenzyme model systems

To understand the biological transport and utilization of dioxygen at the level of mechanistic chemistry it has been necessary to synthesize small molecules (active site sections) which carry out the binding or catalytic function. We have prepared iron porphyrin compounds which mimic the dioxygen binding to myoglobin and the oxidations catalyzed by peroxidases and related enzymes. Using a combination of structural and environmental perturbations and millisecond to subpicosecond laser photolysis methods of following bimolecular or cage reactions we have established many of the factors which control dioxygen binding to heme proteins as well as details of cage processes in ligand binding to small molecules. Additionally, effective catalysts for biomimetic oxidations have been developed and studied by kinetic methods. Studies of the effects of catalyst, oxidant and substrate structure and of environment on the rates of these catalytic processes have allowed mechanisms of epoxidation, hydroxylation and biomimetic suicide labelling to be determined. INTRODUCTION Dioxygen is transported and used in many biological systems through chemical reactions of heme proteins and other metalloproteins (ref. 1). The first protein crystal structures revealed that the active site for transport is an iron@) protoporphyrin complex buried in the protein and attached through the iron to a protein base, often an imidazole of a histidine group (ref. 2). Later biochemical studies revealed that this same site carries out a variety of reactions, e.g. the reduction of hydrogen peroxide (ref. 3). MODEL SYSTEMS FOR BIOLOGICAL OXYGEN TRANSPORT Efforts to synthesize small molecule assemblies which duplicate the structure as well as the reactions of the heme proteins have taken two directions: preparation of stable five coordinated iron(I1) porphyrin (heme) species and, simulation of the pocket in the protein with a large protecting group over or around the iron (ref. 4). Our approach to these two objectives involved the synthesis of a protected porphyrin, a "cyclophane porphyrin" for the pocket (ref. 5) and covalent attachment of an imidazole to analogues of protoheme, systems we have called "chelated hemes" (ref. 6). Dioxygen binding to hemes was first observed at low temperature (ref. 6) but in subsequent studies both steric protection, which prevents bimolecular self oxidation of iron(I1) (e.g. picket fence hemes (ref. 7)) or capped hemes (ref. 8) and fast kinetic methods (ref. 9), in which kinetics and equilibria are easily measured before oxidation sets in, were used to study dioxygen binding. There are now many examples of superstructured hemes which form stable complexes with dioxygen at room temperature (ref. 4). Extensive studies (ref. 4, 10) in these several kinds of model systems have revealed the principal factors which can control dioxygen (and other ligand) binding. Table 1 summarizes the effects of solvent polarity, electron donation to the heme or the fifth ligand, steric hindrance, and hydrogen bonding. Table 1. Effects of Structure and Environment on the Kinetic and Equilibrium Constants for Binding 02 and CO. Environmental Change Resulting Effect on Constantsa co 0 2 on off & a m & 1. Electron Donating Group N N N N D I 2. More Polar Solvent or Nearby Group N i d N D I 3. Hydrogen Bonding to Ligand N N N N d i 4. Increased Steric Hindrance D N D D N D a On = association rate, G; Off = dissociation rate, k i ; Eq = equilibrium constant, K: ; N = no effect, D = large decrease, d = small decrease, I = large increase, i = small increase. This table suggests that, in the relative binding of CO or 0 2 , the differentiation is accomplished by polar effects and not by steric hindrance. Although this conclusion has been subject to some controversy, due in part to the finding that both CO and 0 2 form bent complexes in proteins (ref. 11) but only 9 forms a bent complex with unhindered models (ref. 12), it seems to be generally accepted (ref. 13). The fast kinetic studies, described below also address this point. 265