What External Perturbations Influence the Electronic Properties of Catalase Compound I?

We have performed density functional theory calculations on an active-site model of catalase compound I and studied the responses of the catalytic center to external perturbations. Thus, in the gas phase, compound I has close-lying doublet and quartet spin states with three unpaired electrons:  two residing in π*FeO orbitals and the third on the heme. The addition of a dielectric constant to the model changes the doublet−quartet energy ordering but keeps the same electronic configuration. By contrast, the addition of an external electric field along one of the principal axes of the system can change the doublet−quartet energy splitting by as much as 6 kcal mol-1 in favor of either the quartet or the doublet spin state. This sensitivity is much stronger than the effect obtained for iron heme models with thiolate or imidazole axial ligands. Moreover, an external electric field is able to change the electronic system from a heme-based radical [FeO(Por•+)OTyr-] to a tyrosinate radical [FeO(Por)OTyr•]. This again shows that oxo−iron heme systems are chameleonic species that are influenced by external perturbations and change their character and catalytic properties depending on the local environment

What Affects the Quartet−Doublet Energy Splitting in Peroxidase Enzymes?

Density functional theory calculations have been performed on the active species (Compound I) of cytochrome c peroxidase (CcP) and ascorbate peroxidase (APX) models. We have calculated a large model containing oxo−iron porphyrin plus a hydrogen-bonded network of the axial bound imidazole ligand connected to an acetic acid and an indole group, which mimic the His175, Asp235, and Trp191 amino acids in cytochrome c peroxidase. Our optimized geometries are in good agreement with X-ray and crystallographic structures and give an electronic ground state in agreement with EPR and ENDOR results. We show that the quartet−doublet state ordering and the charge distribution within the model are dependent on small external perturbations. In particular, a single point charge at a distance of 8.7 Å is shown to cause delocalization of the charge and radical characters within the model, thereby creating either a pure porphyrin cation radical state or a tryptophan cation radical state. Thus, our calculations show that small external perturbations are sufficient to change the electronic state of the active species and subsequently its catalytic properties. Similar effects are possible with the addition of an electric field strength along a specific coordination axis of the system. The differences between the electronic ground states of CcP and APX Cpd I are analyzed on the basis of external perturbations

Preferential Hydroxylation over Epoxidation Catalysis by a Horseradish Peroxidase Mutant: A Cytochrome P450 Mimic

Density functional theory calculations are presented on the catalytic properties of a horseradish peroxidase mutant whereby the axial nitrogen atom is replaced by phosphorus. This mutant has never been studied experimentally and only one theoretical report on this system is known (de Visser, S. P. J. Phys. Chem. B 2006, 110, 20759−20761). Thus, a one-atom substitution in horseradish peroxidase changes the properties of the catalytic center of the enzyme to more cytochrome P450-type qualities. In particular, the phosphorus-substituted horseradish peroxidase mutant reacts with substrates via a unique reactivity pattern, whereby alkanes are regioselectively hydroxylated even in the presence of a double bond. Reaction barriers of propene epoxidation and hydroxylation are almost identical to ones observed for a cytochrome P450 catalyst and significantly higher than those obtained for a horseradish peroxidase catalyst. It is shown that the regioselectivity difference is entropy and thermally driven and that the electron-transfer processes that occur during the reaction mechanism follow cytochrome P450-type patterns in the hydroxylation reaction

Propene Activation by the Oxo-Iron Active Species of Taurine/α-Ketoglutarate Dioxygenase (TauD) Enzyme. How Does the Catalysis Compare to Heme-Enzymes?

Density functional calculations on the oxygenation reaction of propene by a model for taurine/α-ketoglutarate dioxygenase (TauD) enzyme are presented. The oxo-iron active species of TauD is shown to be a powerful and aggressive oxidant, which is able to hydroxylate C−H bonds and epoxidize CC bonds with low barriers. In the case of propene oxygenation, the hydroxylation and epoxidation mechanisms are competitive on a dominant quintet spin state surface. We have compared the mechanism and thermodynamics of TauD with oxo-iron heme catalysts, such as the cytochromes P450, and found some critical differences. The TauD model is found to be much more reactive toward oxygenation of substrates than oxo-iron complexes in a heme environment with much lower reaction barriers. We have analyzed this and assigned this to the strength of the O−H bond formed after hydrogen abstraction from a substrate, which is at least 10 kcal mol-1 stronger in five-coordinated oxo-iron nonheme complexes than in six-coordinated oxo-iron heme complexes. Since, the metal in TauD enzymes is five-coordinated, whereas in heme-enzymes it is six-coordinated there are some critical differences in the valence molecular orbitals. Thus, in oxo-iron heme catalysts one of the antibonding π* orbitals is replaced by a low-lying nonbonding δ orbital resulting in a lower overall spin state. Moreover, heme-enzymes have an extra oxidation equivalent located on the heme, which is missing in non-heme oxo-iron catalysts. As a result, the oxo-iron species of TauD reacts via single-state reactivity on a dominant quintet spin state surface, whereas oxo-iron heme catalysts react via two-state reactivity on competing doublet and quartet spin states

Can the Replacement of a Single Atom in the Enzyme Horseradish Peroxidase Convert It into a Monoxygenase? A Density Functional Study

Density functional calculations on horseradish peroxidase mutants are presented, whereby one or two of the nitrogen atoms of the axial histidine ligand have been replaced by phosphorus atoms. Our calculations show that phosphorus entices a push effect on the oxoiron group, whereas a histidine side chain withdraws electrons. As a result, we predict that a phosphorus-substituted histidine ligand will convert the active form of a peroxidase into a monoxygenase. This subsitution may be useful for the bioengineering of commercially exploitable enzymes

van der Waals Equation of State Revisited: Importance of the Dispersion Correction

One of the most basic equations of state describing nonideal gases and liquids is the van der Waals equation of state, and as a consequence, it is generally taught in most first year undergraduate chemistry courses. In this work, we show that the constants a and b in the van der Waals equation of state are linearly proportional to the polarizability volume of the molecules in a gas or liquid. Using this information, a new thermodynamic one-parameter equation of state is derived that contains experimentally measurable variables and physics constants only. This is the first equation of state apart from the Ideal Gas Law that contains experimentally measurable variables and physics constants only, and as such, it may be a very useful and practical equation for the description of dilute gases and liquids. The modified van der Waals equation of state describes pV as the sum of repulsive and attractive intermolecular interaction energies that are represented by an exponential repulsion function between the electron clouds of the molecules and a London dispersion component, respectively. The newly derived equation of state is tested against experimental data for several gas and liquid examples, and the agreement is satisfactory. The description of the equation of state as a one-parameter function also has implications on other thermodynamic functions, such as critical parameters, virial coefficients, and isothermal compressibilities. Using our modified van der Waals equation of state, we show that all of these properties are a function of the molecular polarizability volume. Correlations of experimental data confirm the derived proportionalities

What Factors Influence the Ratio of CH Hydroxylation versus CC Epoxidation by a Nonheme Cytochrome P450 Biomimetic?

Density functional calculations on a nonheme biomimetic (FeO(TMCS)+) have been performed and its catalytic properties versus propene investigated. Our studies show that this catalyst is able to chemoselectively hydroxylate CH bonds even in the presence of CC double bonds. This phenomenon has been analyzed and found to occur due to Pauli repusions between protons on the TMCS ligand with protons attached to the approaching substrate. The geometries of the rate determining transition states indicate that the steric hindrance is larger in the epoxidation transition states than in the hydroxylation ones with much shorter distances; hence the hydroxylation pathway is favored over the epoxidation. Although, the reactant experiences close lying triplet and quintet spin states, the dominant reaction mechanism takes place on the quintet spin state surface; i.e., FeO(TMCS)+ reacts via single-state reactivity. Our calculations show that this spin state selectivity is the result of geometric orientation of the transition state structures, whereby the triplet ones are destabilized by electrostatic repulsions between the substrate and the ligand while the quintet spin transition states are aligned along the ideal axis. The reactivity patterns and geometries are compared with oxoiron species of dioxygenase and monoxygenase enzymes. Thus, FeO(TMCS)+ shows some similarities with P450 enzyme reactivity:  it chemoselectively hydroxylates CH bonds even in the presence of a CC double bond and therefore is an acceptable P450 biomimetic. However, the absolute barriers of substrate oxidation by FeO(TMCS)+ are higher than the ones obtained with heme enzymes, but the chemoselectivity is lesser affected by external perturbations such as hydrogen bonding of a methanol molecule toward the thiolate sulfur or a dielectric constant. This is the first oxoiron complex whereby we calculated a chemoselective hydroxylation over epoxidation in the gas phase

Trends in Substrate Hydroxylation Reactions by Heme and Nonheme Iron(IV)-Oxo Oxidants Give Correlations between Intrinsic Properties of the Oxidant with Barrier Height

Iron(IV)-oxo species have been characterized in several nonheme enzymes and biomimetic systems and are efficient oxidants of aliphatic hydroxylation reactions. However, there appears to be a large variation in substrate hydroxylation ability by different iron(IV)-oxo oxidants due to the effect of the ligands bound to the metal. In this work, we have studied these indirect effects of ligands perpendicular (cis or equatorial) and opposite (trans or axial) to the iron(IV)-oxo group in heme and nonheme oxidants on the oxygenation capability of the oxidant. To this end, we have done a series of density functional theory calculations on the hydrogen atom abstraction of propene by a range of different iron(IV)-oxo oxidants that include heme and nonheme iron(IV)-oxo oxidants. We show that the hydrogen atom abstraction barrier of substrate hydroxylation correlates linearly with the strength of the Fe(III)O−H bond that is formed, i.e., BDEOH, and that this value ranges by at least 20 kcal mol−1 dependent on the cis- and trans-ligands attached to the metal. Thus, our studies show that ligands bound to the metal are noninnocent and influence the catalytic properties of the metal-oxo group dramatically due to involvement into the high-lying occupied and virtual orbitals. A general valence bond curve crossing model is set up that explains how the rate constant of hydrogen atom abstraction is proportional to the difference in energy of the C−H bond of the substrate that is broken and the O−H bond of the Fe(III)O−H complex that is formed, i.e., proportional to BDECH − BDEOH or the reaction enthalpy. In addition, we show a correlation between the polarizability change and barrier height for the hydrogen atom abstraction reaction

A Proton-Shuttle Mechanism Mediated by the Porphyrin in Benzene Hydroxylation by Cytochrome P450 Enzymes

Benzene hydroxylation is a fundamental process in chemical catalysis. In nature, this reaction is catalyzed by the enzyme cytochrome P450 via oxygen transfer in a still debated mechanism of considerable complexity. The paper uses hybrid density functional calculations to elucidate the mechanisms by which benzene is converted to phenol, benzene oxide, and ketone, by the active species of the enzyme, the high-valent iron−oxo porphyrin species. The effects of the protein polarity and hydrogen-bonding donation to the active species are mimicked, as before (Ogliaro, F.; Cohen, S.; de Visser, S. P.; Shaik, S. J. Am. Chem. Soc. 2000, 122, 12892−12893). It is verified that the reaction does not proceed either by hydrogen abstraction or by initial electron transfer (Ortiz de Montellano, P. R. In Cytochrome P450:  Structure, Mechanism and Biochemistry, 2nd ed.; Ortiz de Montellano, P. R., Ed.; Plenum Press:  New York, 1995; Chapter 8, pp 245−303). In accord with the latest experimental conclusions, the theoretical calculations show that the reactivity is an interplay of electrophilic and radicalar pathways, which involve an initial attack on the π-system of the benzene to produce σ-complexes (Korzekwa, K. R.; Swinney, D. C.; Trager, W. T. Biochemistry 1989, 28, 9019−9027). The dominant reaction channel is electrophilic and proceeds via the cationic σ-complex, 23, that involves an internal ion pair made from a cationic benzene moiety and an anionic iron porphyrin. The minor channel proceeds by intermediacy of the radical σ-complex, 22, in which the benzene moiety is radicalar and the iron−porphyrin moiety is neutral. Ring closure in these intermediates produces the benzene oxide product (24), which does not rearrange to phenol (27) or cyclohexenone (26). While such a rearrangement can occur post-enzymatically under physiological conditions by acid catalysis, the computations reveal a novel mechanism whereby the active species of the enzyme catalyzes directly the production of phenol and cyclohexenone. This enzymatic mechanism involves proton shuttles mediated by the porphyrin ring through the N-protonated intermediate, 25, which relays the proton either to the oxygen atom to form phenol (27) or to the ortho-carbon atom to produce cyclohexenone product (26). The formation of the phenol via this proton-shuttle mechanism will be competitive with the nonenzymatic conversion of benzene oxide to phenol by external acid catalysis. With the assumption that 25 is not fully thermalized, this novel mechanism would account also for the observation that there is a partial skeletal retention of the original hydrogen of the activated C−H bond, due to migration of the hydrogen from the site of hydroxylation to the adjacent carbon (so-called “NIH shift” (Jerina, D. M.; Daly, J. W. Science 1974, 185, 573−582)). Thus, in general, the computationally discovered mechanism of a porphyrin proton shuttle suggests that there is an enzymatic pathway that converts benzene directly to a phenol and ketone, in addition to nonenzymatic production of these species by conversion of arene oxide to phenol and ketone. The potential generality of protonated porphyrin intermediates in P450 chemistry is discussed in the light of the H/D exchange observed during some olefin epoxidation reactions (Groves, J. T.; Avaria-Neisser, G. E.; Fish, K. M.; Imachi, M.; Kuczkowski, R. J. Am. Chem. Soc. 1986, 108, 3837−3838) and the general observation of heme alkylation products (Kunze, K. L.; Mangold, B. L. K.; Wheeler, C.; Beilan, H. S.; Ortiz de Montellano, P. R. J. Biol. Chem. 1983, 258, 4202−4207). The competition, similarities, and differences between benzene oxidation viz. olefin epoxidation and alkanyl C−H hydroxylation are discussed, and comparison is made with relevant experimental and computational data. The dominance of low-spin reactivity in benzene hydroxylation viz. two-state reactivity (Shaik, S.; de Visser, S. P.; Ogliaro, F.; Schwarz, H.; Schröder, D. Curr. Opin. Chem. Biol. 2002, 6, 556−567) in olefin epoxidation and alkane hydroxylation is traced to the loss of benzene resonance energy during the bond activation step

Carbon Dioxide: A Waste Product in the Catalytic Cycle of α-Ketoglutarate Dependent Halogenases Prevents the Formation of Hydroxylated By-Products

We present the first density functional theory study on α-ketoglutarate dependent halogenases and focus on the mechanism starting from the iron(IV)-oxo species. The studies show that the high-valent iron(IV)-oxo species reacts with substrates via an initial and rate determining hydrogen abstraction that is characterized by a large kinetic isotope effect (KIE) of 26.7 leading to a radical intermediate. This KIE value is in good agreement with experimental data. The reaction proceeds via two-state reactivity patterns on competing quintet and septet spin state surfaces with close lying hydrogen abstraction barriers. However, the septet spin radical intermediate gives very high barriers for hydroxylation and chlorination whereas the barriers on the quintet spin state surface are much lower. The calculations give extra information regarding the nature of the intermediates and a prediction of a new low-energy mechanism starting from the radical intermediate, whereby a waste product from an earlier step in the catalytic cycle (CO2) is recycled and takes the hydroxyl radical away to form bicarbonate via an OH trapping mechanism. As a consequence, this mechanism prevents the occurrence of hydroxylated byproduct and gives a rationale for the sole observance of halogenated products. By contrast, a direct halogenation reaction cannot compete with hydroxylation due to higher reaction barriers. Our findings support experimental work in the field and give a rationale for the lack of hydroxylation products in α-ketoglutarate dependent halogenases