Organic Reactions Catalyzed by Heme Proteins

The monooxygenases [65-70] are characterized by the axial attachment of the heme cofactor to a thiolate functionality from a cysteine residue of the protein matrix. The binding pockets of monooxygenases are usually organized in such a way as to lead the substrate directly to the active FeO subunit of the porphyrin. This 

Most characteristic for the catalytic cycle of heme monooxygenases is the activation of molecular dioxygen to an active FeO moiety and water. Only two out of four oxidation equivalents are thus used for the synthesis of oxygenated products. This fact is often used as a mechanistic possibility of a short-cut, the so-called peroxide shunt, where a 

Other than for the monooxygenases, a two-electron acceptor such as hydrogen peroxide is required as the terminal oxidant for peroxidases. In the so-called resting state the Fe ion is situated in the oxidation state +3. Reaction with hydrogen peroxide proceeds with loss of water and yields a ferryl(IV) radical cation called Compound I, 

As mentioned before, the application of P450-catalyzed monoxygenations in organic synthesis is largely hampered by the insolubility/instability of most of the isolated enzymes of the family and by the complexity of the reduction system. In 

Site-specific saturation at key residues, followed by a high-throughput activity assay, produced the triple mutant F78V/L188Q/A74G, which shows a greatly increased 700-fold efficiency for the hydroxylation of w-octane and a 200-fold increase in the hydroxylation of [i-ionone at the 3-position [114], Stereoselectivity is poor, however, and all diastereomers are present in the products in comparable amounts. A five-fold mutant, the so-called (F87V)LARV, was developed in a related fashion [115] and shown to hydroxylate capric acid, a substrate not attacked by the wild-type enzyme [116]. 

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The selective catalytic epoxidation of alkenes has become the most important reaction catalyzed by heme proteins in organic synthesis. As described above, the monoxygenase activity of a heme peroxidase is restricted to CPO due to the open substrate access of the ferryl subunit for this enzyme. HRP catalyzes epoxidations only after mutagenetic variations, as shown for the substrate trans-P-methylstyrene [234]. An exception of this rule is the regioselective epoxidation of (T.TJ-piperylpiperidide, which is successfully catalyzed by native HRP [265]. 

Many enzymes use redox centers to store and transfer electrons during catalysis. These redox centers can be composed of metals such as iron or cobalt, or organic cofactors such as quinones, amino acid radicals, or flavins. In order to fully appreciate the catalytic mechanisms of these enzymes, it is often necessary to determine the free energy required to reduce or oxidize their protein redox centers. This is called the redox potential. The measurement of enzyme redox potentials can be performed by either direct or indirect electrochemical methods. The type of electrochemistry suitable for a particular protein system is simply dictated by the accessibility of its redox center to the electrode surface. Because most reactions catalyzed by enzymes occur within hydrophobic pockets of the protein, the redox sites are often far from the surface of the protein. Unless an electron transfer path exists from the protein surface to the redox center, it is not feasible to use direct electrochemistry to measure the redox potential. Since only a few enzymes (most notably certain heme-containing enzymes) have such electron transferring paths and... 

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