Setting an Upper Limit on the Myoglobin Iron(IV)Hydroxide p<i>K</i>_a: Insight into Axial Ligand Tuning in Heme Protein Catalysis

To provide insight into the iron­(IV)­hydroxide p<i>K</i>_a of histidine ligated heme proteins, we have probed the active site of myoglobin compound II over the pH range of 3.9–9.5, using EXAFS, Mössbauer, and resonance Raman spectroscopies. We find no indication of ferryl protonation over this pH range, allowing us to set an upper limit of 2.7 on the iron­(IV)­hydroxide p<i>K</i>_a in myoglobin. Together with the recent determination of an iron­(IV)­hydroxide p<i>K</i>_a ∼ 12 in the thiolate-ligated heme enzyme cytochrome P450, this result provides insight into Nature’s ability to tune catalytic function through its choice of axial ligand

Direct Observation of Oxygen Rebound with an Iron-Hydroxide Complex

The rebound mechanism for alkane hydroxylation was invoked over 40 years ago to help explain reactivity patterns in cytochrome P450, and subsequently has been used to provide insight into a range of biological and synthetic systems. Efforts to model the rebound reaction in a synthetic system have been unsuccessful, in part because of the challenge in preparing a suitable metal-hydroxide complex at the correct oxidation level. Herein we report the synthesis of such a complex. The reaction of this species with a series of substituted radicals allows for the direct interrogation of the rebound process, providing insight into this uniformly invoked, but previously unobserved process

Direct Observation of Oxygen Rebound with an Iron-Hydroxide Complex

The rebound mechanism for alkane hydroxylation was invoked over 40 years ago to help explain reactivity patterns in cytochrome P450, and subsequently has been used to provide insight into a range of biological and synthetic systems. Efforts to model the rebound reaction in a synthetic system have been unsuccessful, in part because of the challenge in preparing a suitable metal-hydroxide complex at the correct oxidation level. Herein we report the synthesis of such a complex. The reaction of this species with a series of substituted radicals allows for the direct interrogation of the rebound process, providing insight into this uniformly invoked, but previously unobserved process

Spectroscopic Investigations of Catalase Compound II: Characterization of an Iron(IV) Hydroxide Intermediate in a Non-thiolate-Ligated Heme Enzyme

We report on the protonation state of <i>Helicobacter pylori</i> catalase compound II. UV/visible, Mössbauer, and X-ray absorption spectroscopies have been used to examine the intermediate from pH 5 to 14. We have determined that HPC-II exists in an iron­(IV) hydroxide state up to pH 11. Above this pH, the iron­(IV) hydroxide complex transitions to a new species (p<i>K</i>_a = 13.1) with Mössbauer parameters that are indicative of an iron­(IV)-oxo intermediate. Recently, we discussed a role for an elevated compound II p<i>K</i>_a in diminishing the compound I reduction potential. This has the effect of shifting the thermodynamic landscape toward the two-electron chemistry that is critical for catalase function. In catalase, a diminished potential would increase the selectivity for peroxide disproportionation over off-pathway one-electron chemistry, reducing the buildup of the inactive compound II state and reducing the need for energetically expensive electron donor molecules

Oxygen-Atom Transfer Reactivity of Axially Ligated Mn(V)–Oxo Complexes: Evidence for Enhanced Electrophilic and Nucleophilic Pathways

Addition of anionic donors to the manganese­(V)–oxo corrolazine complex Mn^V(O)­(TBP₈Cz) has a dramatic influence on oxygen-atom transfer (OAT) reactivity with thioether substrates. The six-coordinate anionic [Mn^V(O)­(TBP₈Cz)­(X)]⁻ complexes (X = F^–, N₃^–, OCN^–) exhibit a ∼5 cm^–1 downshift of the Mn–O vibrational mode relative to the parent Mn^V(O)­(TBP₈Cz) complex as seen by resonance Raman spectroscopy. Product analysis shows that the oxidation of thioether substrates gives sulfoxide product, consistent with single OAT. A wide range of OAT reactivity is seen for the different axial ligands, with the following trend determined from a comparison of their second-order rate constants for sulfoxidation: five-coordinate ≈ thiocyanate ≈ nitrate < cyanate < azide < fluoride ≪ cyanide. This trend correlates with DFT calculations on the binding of the axial donors to the parent Mn^V(O)­(TBP₈Cz) complex. A Hammett study was performed with <i>p</i>-X-C₆H₄SCH₃ derivatives and [Mn^V(O)­(TBP₈Cz)­(X)]⁻ (X = CN^– or F^–) as the oxidant, and unusual “V-shaped” Hammett plots were obtained. These results are rationalized based upon a change in mechanism that hinges on the ability of the [Mn^V(O)­(TBP₈Cz)­(X)]⁻ complexes to function as either an electrophilic or weak nucleophilic oxidant depending upon the nature of the <i>para</i>-X substituents. For comparison, the one-electron-oxidized cationic Mn^V(O)­(TBP₈Cz^•+) complex yielded a linear Hammett relationship for all substrates (ρ = −1.40), consistent with a straightforward electrophilic mechanism. This study provides new, fundamental insights regarding the influence of axial donors on high-valent Mn^V(O) porphyrinoid complexes

O₂-Evolving Chlorite Dismutase as a Tool for Studying O₂-Utilizing Enzymes

The direct interrogation of fleeting intermediates by rapid-mixing kinetic methods has significantly advanced our understanding of enzymes that utilize dioxygen. The gas’s modest aqueous solubility (<2 mM at 1 atm) presents a technical challenge to this approach, because it limits the rate of formation and extent of accumulation of intermediates. This challenge can be overcome by use of the heme enzyme chlorite dismutase (Cld) for the rapid, <i>in situ</i> generation of O₂ at concentrations far exceeding 2 mM. This method was used to define the O₂ concentration dependence of the reaction of the class Ic ribonucleotide reductase (RNR) from <i>Chlamydia trachomatis</i>, in which the enzyme’s Mn^IV/Fe^III cofactor forms from a Mn^II/Fe^II complex and O₂ via a Mn^IV/Fe^IV intermediate, at effective O₂ concentrations as high as ∼10 mM. With a more soluble receptor, myoglobin, an O₂ adduct accumulated to a concentration of >6 mM in <15 ms. Finally, the C–H-bond-cleaving Fe^IV–oxo complex, <b>J</b>, in taurine:α-ketoglutarate dioxygenase and superoxo–Fe₂^III/III complex, <b>G</b>, in <i>myo</i>-inositol oxygenase, and the tyrosyl-radical-generating Fe₂^III/IV intermediate, <b>X</b>, in <i>Escherichia coli</i> RNR, were all accumulated to yields more than twice those previously attained. This means of <i>in situ</i> O₂ evolution permits a >5 mM “pulse” of O₂ to be generated in <1 ms at the easily accessible Cld concentration of 50 μM. It should therefore significantly extend the range of kinetic and spectroscopic experiments that can routinely be undertaken in the study of these enzymes and could also facilitate resolution of mechanistic pathways in cases of either sluggish or thermodynamically unfavorable O₂ addition steps

Oxygen-atom transfer reactivity of axially ligated Mn(V)−oxo complexes: Evidence for enhanced electrophilic and nucleophilic pathways

Addition of anionic donors to the manganese(V)−oxo corrolazine complex MnV(O)(TBP8Cz) has a dramatic influence on oxygen-atom transfer (OAT) reactivity with thioether substrates. The sixcoordinate anionic [MnV(O)(TBP8Cz)(X)]− complexes (X = F−, N3−, OCN−) exhibit a ∼5 cm−1 downshift of the Mn−O vibrational mode relative to the parent MnV(O)(TBP8Cz) complex as seen by resonance Raman spectroscopy. Product analysis shows that the oxidation of thioether substrates gives sulfoxide product, consistent with single OAT. A wide range of OAT reactivity is seen for the different axial ligands, with the following trend determined from a comparison of their second-order rate constants for sulfoxidation: five-coordinate ≈ thiocyanate ≈ nitrate < cyanate < azide < fluoride ≪ cyanide. This trend correlates with DFT calculations on the binding of the axial donors to the parent MnV(O)(TBP8Cz) complex. A Hammett study was performed with p-X-C6H4SCH3 derivatives and [MnV(O)(TBP8Cz)(X)]−(X = CN− or F−) as the oxidant, and unusual “V-shaped” Hammett plots were obtained. These results are rationalized based upon a change in mechanism that hinges on the ability of the [MnV(O)(TBP8Cz)(X)]− complexes to function as either an electrophilic or weak nucleophilic oxidant depending upon the nature of the para-X substituents. For comparison, the oneelectron-oxidized cationic MnV(O)(TBP8Cz•+) complex yielded a linear Hammett relationship for all substrates (ρ = −1.40), consistent with a straightforward electrophilic mechanism. This study provides new, fundamental insights regarding the influence of axial donors on high-valent MnV(O) porphyrinoid complexes