Nonheme oxo-iron(IV) intermediates form an oxyl radical upon approaching the C–H bond activation transition state

Shengfa Ye and Frank Neese1

Lehrstuhl für Theoretische Chemie, Universität Bonn, Wegelerstrasse 12, D-53115 Bonn, Germany

Edited* by Brian M. Hoffman, Northwestern University, Evanston, IL, and approved December 5, 2010 (received for review June 29, 2010)

Oxo-iron(IV) species are implicated as key intermediates in the mately 100 times faster than the corresponding mononuclear IS catalytic cycles of heme and nonheme oxygen activating iron oxo-iron(IV) complex (8). This experimental finding is in accord enzymes that selectively functionalize aliphatic C–H bonds. Ferryl with the theoretical prediction that quintet ferryl species are complexes can exist in either quintet or triplet ground states. more aggressive oxidants than the corresponding triplet counter- Density functional theory calculations predict that the quintet parts (9, 10, 11). However, there is a long-term debate on how to oxo-iron(IV) species is more reactive toward C–H bond activation rationalize the differential reactivity between quintet and triplet than its corresponding triplet partner, however; the available oxo-iron(IV) species. De Visser, Shaik, and coworkers argued experimental data on model complexes suggests that both spin that the enhanced exchange interaction upon approaching the multiplicities display comparable reactivities. To clarify this ambigu- transition state (TS) on the quintet surface flattens the potential ity, a detailed electronic structure analysis of hydroxylation energy surface (PES) and hence lowers the barrier (12, 13). Baer- by an oxo-iron(IV) species on different spin-state potential energy ends, Solomon, and coworkers proposed that the greater degree surfaces is performed. According to our results, the lengthening of exchange stabilization in the HS ferryl reactant relative to the of the Fe–oxo bond in ferryl reactants, which is the part of the IS analogues significantly stabilizes the Fe-dz2 based σ-antibond- reaction coordinate for H-atom abstraction, leads to the formation ing molecular orbital (MO) (14–18). Therefore, this orbital is of oxyl-iron(III) species that then perform actual C–H bond activa- able to serve as the acceptor on the quintet surface, tion. The differential reactivity stems from the fact that the two whereas the energy of the corresponding MO in the triplet reac- spin states have different requirements for the optimal angle at tant is too high to be an electron acceptor. Thus, the lower energy þ which the substrate should approach the ðFeOÞ2 core because π-antibonding MOs that mainly consist of the Fe-dxz∕yz and the distinct electron acceptor orbitals are employed on the two sur- O-px∕y fragment orbitals act as electron acceptors on the triplet faces. The H-atom abstraction on the quintet surface favors the surface (17). As a consequence, a σ-pathway is often suggested to “σ-pathway” that requires an essentially linear attack; by contrast be operative on the quintet surface, whereas the triplet reaction a “π-channel” is operative on the triplet surface that leads to an proceeds via a π-channel (18). However, under certain circum- ideal attack angle near 90°. However, the latter is not possible stances quintet π- and triplet σ-pathways were also reported in due to steric crowding; thus, the attenuated orbital interaction the process of C–H bond activation (19, 20). Recent studies and the unavoidably increased Pauli repulsion result in the lower on the reaction of alkane hydroxylation by ferryl model com- reactivity of the triplet oxo-iron(IV) complexes. pounds demonstrated that the reaction can take place through both the σ- and the π-pathways on the quintet and triplet surfaces 5 5 3 3 density functional calculation ∣ nonheme iron ∣ reaction mechanism with barrier heights displaying the order σ > π ≈ π > σ (21). The results indicated that quintet oxo-iron(IV) species are not – xo-iron(IV) intermediates have attracted much interest in always more reactive toward C H than the corre- bioinorganic because they are implicated as key sponding triplet species and confirmed that of all possible path- O σ intermediates in the catalytic cycles of heme and nonheme oxy- ways the quintet -pathway is the most effective one. More gen activating iron enzymes that selectively functionalize unacti- importantly, a careful analysis of the electronic structure changes vated C–H bonds (1). Detailed experimental and theoretical along the quintet pathway revealed that the reaction requires a studies on the hydroxylation of saturated hydrocarbons by ferryl preparatory stage in which an oxyl-iron(III) species is formed en species in heme systems, foremost cytochrome P450, have been route to the TS that then functions as the active species in the – performed (2). On the other hand a number of nonheme enzymes actual C H bond activation process (22). Thus far all interpreta- – are able to activate molecular dioxygen to modify alkane or tions about which spin multiplicity is more reactive toward C H “ ” arene substrates as well. So far, nonheme ferryl species have bond activation have been based on the notion that the real been spectroscopically characterized in four mononuclear iron oxidant is oxo-iron(IV) rather than oxyl-iron(III). Therefore, enzymes and were found to feature high-spin (HS) (S ¼ 2) elec- to address this question one needs to understand how the real tronic ground state configurations (3). In parallel, a wide range of oxyl-ferric reactant is formed and how it then reacts with the sub- synthetic FeðIVÞ¼O complexes were synthesized and character- strate on the quintet and triplet surfaces, respectively. ized (4). In almost all cases they contain intermediate-spin (IS) In the present contribution we address the above formulated (S ¼ 1) rather than HS ferryl centers (4). The only exceptions are problem through an analysis of the electronic structure changes 2þ that occur upon alkane C–H bond hydroxylation by the taurine: ½FeðIVÞðOÞðH2OÞ5 (5) and the recently reported model com- 2þ plex ½FeðIVÞðOÞðTMG3trenÞ (TMG3tren ¼ N½CH2CH2N ¼ 2þ CðNMe2Þ2) (6). Presumably because the ðFeOÞ core is shel- Author contributions: S.Y. and F.N. designed research; S.Y. performed research; S.Y. and tered by the sterically bulky supporting ligand in ½FeðIVÞðOÞ F.N. analyzed data; and S.Y. and F.N. wrote the paper. 2þ ðTMG3trenÞ , its reactivity toward C–H bond cleavage is only The authors declare no conflict of interest. comparable with triplet ferryl analogues (7). Recently, an un- *This Direct Submission article had a prearranged editor. masked HS FeðIVÞ¼O unit was identified in a valence-localized 1To whom correspondence should be addressed. E-mail: [email protected]. ð Þ∕ ð Þ open core diiron (Fe III Fe IV ) complex (8). The reaction of This article contains supporting information online at www.pnas.org/lookup/suppl/ the C–H bond cleavage by this complex turned out to be approxi- doi:10.1073/pnas.1008411108/-/DCSupplemental.

1228–1233 ∣ PNAS ∣ January 25, 2011 ∣ vol. 108 ∣ no. 4 www.pnas.org/cgi/doi/10.1073/pnas.1008411108 Downloaded by guest on September 26, 2021 α-ketoglutarate dioxygenase (TauD) oxo-iron(IV) intermediate Table 1. The relative enthalpy, entropy, and free energy for (23) along the septet (hypothetical), quintet, and triplet reaction the key local minima and transition states on the septet, channels. This allows us to gain electronic structure insight into quintet, and triplet surface the differential reactivity of the distinct spin states and draw con- ΔH (kcal∕mol) −TΔS kcal∕mol) ΔG (kcal∕mol) clusions that might have a wider range of applicability. 5R0 0 0 Results 5TS1 14.0 10.6 24.6 5 The energy profile for H-atom abstraction by the TauD oxo-iron IN 0.9 9.0 9.8 7R16.3 −1.3 15.1 (IV) intermediate is shown in Fig. 1. The reaction follows the 7 – TS1 19.1 10.0 29.0 consensus mechanism for the activation of aliphatic C H bonds 7IN 2.7 7.8 10.5 by oxo-iron(IV) compounds (24).The reaction is initiated by the 3R 6.8 2.4 9.2 rate-determining H-atom abstraction reaction via TS1 to yield a 3TS1 28.5 13.7 42.2 hydroxo-iron(III) complex with a nearby alkyl radical intermedi- 3IN 14.1 8.9 23.0 ate (IN). This step is followed by an OH-rebound process leading to the final product ethanol. Interestingly, for H-atom transfer 7 1 the hypothetical septet reaction pathway is calculated to show the The geometry of TS exhibits a slight contraction of the – 7 1 lowest barrier (Fig. 1 and Table 1). However, because the singlet Fe Ooxo bond relative to the reactant (1.89 Å in TS (Table 2) 7 1 product ethanol cannot be formed on the septet surface, the vs. 1.91 Å in the reactant). The electronic structure of TS is best 7 S ¼ 5∕2 reaction on the septet surface stops at intermediate IN. In interpreted as featuring a HS ferric ( Fe ) interacting – σ S ¼ 1∕2 addition, the reaction cannot start on the septet surface, because with a three-center C H-O -radical ( CHO ) in a ferromag- the septet reactant is much higher in energy than the quintet oxo- netic fashion (Fig. 2, Right). In comparison to the electronic struc- iron(IV) reactant. In fact, a low-lying septet state is only possible ture of the reactant, one may envision that during the approach of 7ð Þ2þ for a mononuclear 3d transition metal center if at least one ligand the substrate toward the FeO core, the singly occupied O-px – σ carries an unpaired electron. orbital interacts with the C H -bonding orbital in the substrate σ σ to form a pair of three-centered MOs ( CHO and CHO). Because π Septet. Due to the effective C4v symmetry of the complex, the the electron acceptor for H-atom abstraction is a -bonding or- lowest-energy septet state of the ðFeOÞ2þ core is 7E with the bital (O-px), the septet reaction mechanism employs a π-pathway.

2 3 1 CHEMISTRY orbital occupation pattern ðσðO-pzÞÞ ðπðO-px;yÞÞ ðFe-dxyÞ ðπ After passing through the very low barrier for H-atom abstrac- 2 1 1 S ¼ 5∕2 ðFe-dxz;yzÞÞ ðσ ðFe-dx2−y2ÞÞ ðσ ðFe-dz2ÞÞ (Fig. 2, Left). This cor- tion, the system evolves to a HS Fe(III) ( Fe ) center that is S ¼ 5∕2 ferromagnetically coupled to a substrate alkyl radical (S ¼ 1∕2). responds to a HS ferric center ( Fe ) that is ferromagneti- C S ¼ 1∕2 At this point the first electron transfer has been accomplished. cally coupled to an oxyl-radical ( O ) thereby yielding an overall septet state. Relative to the quintet ground state of the However, ferromagnetic coupling between the HS ferric center 2þ 5 ðFeOÞ core ( A1), the septet state represents an oxo-to-iron and the substrate radical renders the second electron transfer πðO-px;yÞ → σ ðFe-dz2Þ charge transfer excited state that is ac- impossible; thus, the final products cannot be produced on the companied by a spin flip. The orbital occupation pattern implies septet surface. aFe–O bond order of 1 compared to a Fe–O bond order of 2 oxo oxo 5 1 that exists in the quintet ground state. The reduction in the bond Quintet. The most noticeable geometric feature in TS is a fairly – long Fe–O bond in comparison with that in the reactant order results in a significant lengthening of the Fe Ooxo bond oxo 5 2þ 7 2þ 5 1 (1.62 Å in ðFeOÞ vs 1.91 Å in ðFeOÞ ). B3LYP calculations (1.76 Å in TS (Table 2) vs. 1.62 Å in the reactant). As shown 5 1 predict the septet species to be ∼16 kcal∕mol higher in energy in the left column of Fig. 3, the electronic structure of TS is best S ¼ 5∕2 than the corresponding quintet ferryl complex. Hence, one can interpreted as a HS Fe(III) ion ( Fe ) antiferromagnetically – – S ¼ 1∕2 safely rule out the possibility that the septet species can be coupled to a three-center C H O radical ( CHO ). Unex- trapped as an intermediate in enzymes or model systems. How- pectedly, the Fe-dz2 based MO is the bonding combination ever, among the three alternative spin-state surfaces, the septet between the Fe-dz2 and the O-pz fragment orbitals rather than species is the most reactive toward H-atom abstraction. Thus, a the antibonding combination that would be expected from the discussion of its electronic structure still provides valuable clues electronic structure of the quintet oxo-iron(IV) reactant. As dis- to understanding the reaction mechanism on the other two cussed elsewhere (22), this signifies that dramatic changes in the spin-state surfaces that will be investigated below. electronic structure of the quintet ferryl species must have oc- – 5 1 curred as the Fe Ooxo bond is lengthened to approach TS . The evolution of the electronic structure of the quintet oxo- iron(IV) complex was investigated more closely through a relaxed – surface scan at a series of fixed Fe Ooxo bond distances (Fig. 3, – Right). When the Fe Ooxo bond is elongated to 1.80 Å similar to 5 that found in TS1, the σ-bonding MO that is mainly of O-pz char- acter at the equilibrium geometry splits into a spin-coupled pair. One can identify that the MO in the spin-up manifold becomes Fe-dz2 based (Fe 61% vs O 36%) and that the MO in the spin- down set acquires more O-pz character (Fe 24% vs O 68%). Thus,

Table 2. The important geometric features of the key local minima and transition states on the septet, quintet, and triplet surfaces 7TS1 7IN 5TS1 5IN 3TS1 3IN O–Fe (Å) 1.890 1.847 1.765 1.838 1.756 1.793 O–H (Å) 1.279 0.981 1.224 0.986 1.164 0.987 C–H (Å) 1.262 2.368 1.289 2.231 1.388 2.442 Fe–O–H (degree) 113.1 123.0 165.0 122.0 119.2 107.0 Fig. 1. The energy profile of H-atom abstraction from ethane by the TauD O–H–C (degree) 178.7 — 177.9 — 173.5 — oxo-iron(IV) intermediate (B3LYP/def2-TZVPP).

Ye and Neese PNAS ∣ January 25, 2011 ∣ vol. 108 ∣ no. 4 ∣ 1229 Downloaded by guest on September 26, 2021 Fig. 2. Schematic MO diagrams for the septet reactant (Left) and 7TS1 (Right).

– α the elongation of the Fe Ooxo bond leads to a crossing of the The oxyl oxygen may more easily accept an -electron from ground state [best described as oxo-iron(IV)] with a state that the substrate C–H bond. In this electron transfer process the S ¼ 5∕2 is best characterized as a HS ferric center ( Fe ) antiferro- bonding nature of the Fe-dz2 based MO is retained and the inter- S ¼ 1∕2 magnetically coupled to an oxyl-radical ( O ). Accordingly, action of the singly occupied O-pz orbital with the electron donor σ σ the lowest unoccupied molecular orbital in the spin-up manifold affords a pair of three-centered MOs ( CHO and CHO). Thus, that is the Fe-dz2 based σ*-orbital at the equilibrium geometry one may visualize the entire C–H bond activation process as con- – evolves to an O-pz based orbital (Fig. 3, Right). As the Fe Ooxo sisting of a preparatory stage [the formation of the oxyl-iron(III) bond is lengthened, the energy of this orbital is significantly species] and the actual H-atom abstraction that is accomplished −2 4 – −3 0 – lowered from . eV (Fe Ooxo 1.65 Å) to . eV (Fe Ooxo by the highly reactive oxyl-radical. It is important to point out that 1.80 Å). Moreover, the O-pz based orbital overlaps more effi- the oxyl-iron(III) species does not represent a minimum on the σ ciently with the electron donor C-H than the Fe-dz2 based orbital PES and hence is not a true reaction intermediate. This way of that has limited O-pz character. Therefore, the oxyl species that is looking at the reaction is simply an approach to rationalize the generated possesses higher electrophilicity than the oxo-form. electronic structure changes that occur en route to the H-atom

5 1 – Fig. 3. Schematic MO diagrams for TS (Left) and the evolution of the quintet oxo-iron(IV) species as a function of the Fe Ooxo bond distance (Right).

1230 ∣ www.pnas.org/cgi/doi/10.1073/pnas.1008411108 Ye and Neese Downloaded by guest on September 26, 2021 abstraction TS. As a consequence of this interpretation, the elec- nic structure rearrangement of the triplet ferryl complex occurred 3 tron acceptor for the quintet σ-pathway is best viewed to be the en route to TS1, the Fe-dxz based MO would be the antibonding 3 O-pz based σ*-orbital rather than the Fe-dz2 orbital as proposed combination of the Fe-dxz and O-px fragmental orbitals in TS1. before (10, 12, 14). 5IN and 7IN are essentially energetically de- These results confirm the necessity of the preparatory stage to generate, because they have the same orbital occupation pattern form an oxyl-ferric species that then performs the actual H-atom except for the different spin coupling between the alkyl-radical abstraction. and the HS ferric center. Discussion Triplet. Given the rather high barrier for H-atom abstraction, the Formation of Oxyl-Ferric Species on Quintet and Triplet Surfaces. If triplet channel has no catalytic relevance for C–H bond activa- the interaction between the iron center and the oxo ligand were 5 1 – tion. In analogy to TS , an elongated Fe Ooxo bond was also dominantly ionic, as is the case for classical Werner-type transi- 3 – identified in TS1 (see Table 2). The cleaving C–H bond distance tion metal complexes, lengthening of the Fe Ooxo bond would in 3TS1 is 0.1 Å longer than that in 5TS1. The relative orientation tend to lower the covalency of the Fe-oxo interaction; hence, – of the substrate with respect to the Fe Ooxo core is similar to that the bonding MO that is mainly ligand based would acquire more in 7TS1 but significantly different from that calculated for 5TS1. O-p character, and accordingly the magnitude of the O-p frag- As depicted in the left column of Fig. 4, the electronic structure ment orbital in the antibonding partner would be decreased. As of 3TS1 is best rationalized as consisting of a low-spin (LS) a result, if the Fe-3d based antibonding MOs acted as electron S ¼ 1∕2 – Fe(III) ion ( Fe ) ferromagnetically coupled to a three- acceptors for C H bond activation, the overlap between the – – S ¼ 1∕2 σ center C H O radical ( CHO ). Surprisingly, the Fe-dxz substrate C-H and the O-p fragment orbitals would gradually based MO is the bonding combination of the Fe-dxz and O-px decrease upon approaching the TS. This would be counterpro- fragment orbitals, rather than the expected antibonding combina- ductive. – tion in the triplet oxo-iron(IV) reactant. This suggests that, like in In fact, the lengthening of the Fe Ooxo bond on the quintet the quintet pathway, a preparatory stage may also be needed in surface leads to the homolytic cleavage of the Fe–oxo σ-bond. the triplet reaction channel. In analogy to the elongation of the H–H bond in H2 yielding To probe the changes in the electronic structure of the a singlet diradical, the doubly occupied σ-bonding MO splits into 3ð Þ2þ – FeO core as the Fe Ooxo bond is lengthened, a constrained a spin-coupled pair. Due to the incurred exchange stabilization, – α geometry optimization was performed with a Fe Ooxo bond dis- the -electron resides on the iron center, whereas the oxygen CHEMISTRY tance kept fixed at 1.8 Å. The constraint model is destabilized by atom carries the β-electron. As such, the Fe-dz2 based MO in 8.1 kcal∕mol relative to the triplet reactant. As shown in the right the spin-up manifold is the bonding rather than the antibonding – column of Fig. 4, the elongation of the Fe Ooxo bond results in combination as observed in Fig. 3. In line with this discussion, σ σ qualitative changes in the electronic structure. The description in CHO and CHO the interaction between the Fe-dz2 and S ¼ 1 S ¼ 1∕2 changes from an IS ferryl ( ) to a LS ferric ion ( Fe ) O-pz fragment orbitals is of bonding nature as well. S ¼ 1∕2 – that is ferromagentically coupled to an oxyl-radical ( O ). However, on the triplet surface elongation of the Fe Ooxo At the same time one of the π-bonding MOs becomes domi- bond results in the heterolytic cleavage of the Fe–oxo π-bond. nantly Fe-dxz in character. Accordingly the corresponding π*-MO Due to the fact that the Fe–oxo interaction is predominantly changes to an O-px based orbital. Comparison of the electronic covalent, it follows that these two fragments have comparable structure of the triplet oxyl-ferric species with that of 3TS1 pro- ; thus, one cannot determine a priori eventually vides a coherent picture on how the first electron transfer from whether the O-atom obtains two or the iron center the substrate to the iron center takes place in the triplet reaction acquires two electrons. By inspection of Fig. 4, one can anticipate 3 1 – channel. Upon approach of TS the singly occupied O-px orbital, that upon lengthening of the Fe Ooxo bond on the triplet surface – π formed by the elongation of the Fe Ooxo bond, interacts with the the iron center will obtain the two electrons in the -bonding MO σ C-H bonding orbital thus yielding a pair of three-centered MOs (referred to as Fe-dxz in the right column of Fig. 4), whereas the σ σ π ( CHO and CHO) similar to what is found on the septet surface. O-atom will receive the electron in the -antibonding singly Hence, the triplet reaction mechanism employs a π-pathway using occupied molecular orbital (SOMO) (referred to as O-px). This the O-px based π*-orbital as the electron acceptor. If no electro- results in the observation that the Fe-dxz based MO is of bonding

3 1 – Fig. 4. Schematic MO diagrams for TS (Left) and the constraint model of the triplet oxo-iron(IV) species with the Fe Ooxo bond length at 1.80 Å (Right).

Ye and Neese PNAS ∣ January 25, 2011 ∣ vol. 108 ∣ no. 4 ∣ 1231 Downloaded by guest on September 26, 2021 character and the corresponding antibonding combination is In the quintet σ-mechanism where the O-pz based σ*-orbital σ σ found in CHO and CHO in Fig. 4. If the electron distribution serves as the electron acceptor, the system arranges a vertical patterns were opposite during the heterolysis of the Fe–oxo π- attack of the cleaving C–H bond toward the ðFeOÞ2þ core such π σ bond, the electron acceptor would be the Fe-dxz based *-SOMO. that the overlap between the O-pz and the C–H orbital is max- Upon approaching the TS, the O-px contribution in this SOMO imized. This requires the Fe–O–H arrangement to be approxi- would be decreased and hence so would be the overlap between mately collinear. More importantly, the linear geometry signi- the electron donor and the acceptor. ficantly reduces the Pauli repulsion between the substrate and the oxo-iron(IV) reactant in comparison with the π-pathway Intrinsic Electron Acceptors for H-Atom Abstraction. From the con- (see below) (16). For the π-mechanism the O-px based π*-orbital sensus mechanism one is readily aware that the hydroxylation acts as the electron acceptor. Thus, the optimum Fe–O–H angle of aliphatic C–H bonds by ferryl complexes is a two-electron should be 90° to afford the maximum overlap between the O-px σ σ oxidation process and that the C–H bonding orbital serves as and the C–H orbitals. One readily appreciates that such a strictly the electron donor. However, a number of mechanisms can be horizontal approach of the substrate would result in a large Pauli envisioned that differ in the acceptor orbital on the reactive repulsion. The compromise between the orbital interactions and metal-oxo unit. Pauli repulsion yields a final Fe–O–H angle close to 120° as found Due to the steric hindrance provided by the ligand framework, in 3TS1. This may rationalize the observation that 3TS1 is a the substrate can only interact with the oxo group rather than “later” TS as suggested by a longer C–H bond length identified directly with the iron center. It follows that direct electron trans- in 3TS1 than that in 5TS1. Taken together, the present analysis fer from the substrate to the iron center is very unlikely, and implies that the synergy of the minimal Pauli repulsion and that all possible electron transfer pathways (9) have to pass the favorable orbital interactions render the quintet σ-pathway through the O-atom. In other words, during the reaction the more effective for H-atom abstraction reactions than the triplet O-atom relays the electron from the substrate to the final elec- π-pathway. tron acceptor that is the iron center. Therefore, viable electron Based on the above analysis one may anticipate that the quin- acceptor orbitals should contain significant contributions from tet π-channel may encounter a comparable barrier to that calcu- the O-atom. The Fe-dxy and Fe-dx2−y2 orbitals can be excluded, lated on the triplet surface due to the unavoidably increased Pauli because they are located in the plane perpendicular to the line repulsion and the reduced orbital overlap relative to the quintet of the substrate attack and hence have no significant interactions σ-pathway. This is in agreement with our previous studies on the with the oxo ligand or the substrate. As discussed above, the quin- relative reactivity of all viable reaction channels for alkane hydro- tet and triplet reaction pathways require a preparatory stage to xylation by oxo-iron(IV) complexes (21). For the triplet σ-path- produce an oxyl-ferric species. As a consequence, in the relay way, in the preparatory stage the triplet ferryl species needs to S ¼ 3∕2 pathway the oxo group first delivers one of its electrons to the evolve into an IS ferric ion ( Fe ) that would be antiferro- σ S ¼ 1∕2 iron center and then accepts an electron from the C-H orbital. magnetically coupled to an oxyl radical ( O ). In fact, the Hence, the oxyl-p orbitals act as intrinsic electron acceptors in strength of the Fe–oxo σ-bond is identical irrespective of the spin the actual H-atom abstraction. More importantly, upon ap- multiplicity of the oxo-iron(IV) species (25). The energy penalty proaching 3;5TS1 the oxyl-p based MOs have lower energy and for accomplishing this transformation should be much higher on σ better overlap with the C-H orbital than the Fe-dz2 and Fe-dxz;yz the triplet surface than for the corresponding process on the quin- based orbitals. tet surface because of the substantially reduced exchange stabi- lization generated by three rather than five unpaired electrons in Differential Reactivity of H-Atom Abstraction for Quintet and Triplet the IS ferric center. Hence, the π-pathway is the energetically Ferryl Species. To address the question of why the quintet oxo-iron more favored mechanism on the triplet surface, and our previous (IV) is more reactive toward C–H bond cleavage than the corre- calculations suggested that among the four reaction pathways for sponding triplet species, one may loosely divide the barrier for the H-atom abstraction the triplet σ-pathway has the highest barrier H-atom abstraction process into three contributions: (i) the geo- (21). The observation that the septet channel involves the lowest metric rearrangements, which mainly consists of elongation of the barrier for H-atom abstraction although it employs a π-pathway – ð Þ2þ – Fe Ooxo bond in the FeO core and the cleaving C H bond in may be rationalized as follows: (i) no preparatory stage is needed; the substrate, (ii) Pauli repulsion between the incoming substrate (ii) more importantly, the driving force is quite large because the and the high-valent iron species, and (iii) the orbital interactions final electron acceptor is a bonding rather than an antibonding between the electron donor and acceptor orbitals. Which of these orbital with respect to the Fe–oxo interaction as evidenced by – 7 1 factors is dominant in determining the differential reactivity of a slightly contracted Fe Ooxo bond in TS compared to that the quintet and triplet reaction pathways? in the septet reactant. This is in sharp contrast with what is found – In the preparatory stage the quintet pathway involves the par- on the other two spin-state surfaces where the Fe Ooxo bond is 3;5 tial breaking of the Fe–oxo σ-bond to generate a hole in the O-pz dramatically lengthened in TS1;(iii) it involves reduced Pauli σ π – 7 1 based *-orbital, whereas only half of the -bond is broken in the repulsion caused by the significantly longer Fe Ooxo bond in TS triplet channel (Fig. 3, 4). Moreover, the TSs for H-atom abstrac- (Table 2). 5 1 3 1 – tion ( TS and TS ) exhibit essentially the same Fe Ooxo bond As elaborated above, in terms of the two-electron oxidation of distance. Therefore, one may simply argue that in terms of the by oxo-iron(IV) intermediates several orbitals may act as – Fe Ooxo bond lengthening, the triplet pathway should be energe- electron acceptors. Different electron acceptors require different tically favored. In fact, the energy required for the elongation of optimal attack geometries for the substrate to maximize the – the Fe Ooxo bond in the quintet oxo-iron(IV) intermediate from orbital interactions between the electron donor and the electron its equilibrium geometry (1.65 Å) to 1.80 Å in 5TS1 is 9.4 kcal∕mol, acceptor. Hence, the already inherently complex nature of the whereas the corresponding energy increment is 8.1 kcal∕mol for reactivity of open-shell transition metal complexes is further com- the triplet species. Thus, unexpectedly the quintet channel is only plicated by the fact that different substrate orientations lead to marginally energetically disfavored. This may be ascribed to the different preferred reaction channels depending on the angle much stronger spin-polarization stabilization created by five un- at which the substrate approaches the reactive core. This is clearly paired electrons in the HS ferric center in 5TS1 compared to only a more general feature that is not limited to high-valent oxo-iron oneunpairedelectronintheLSiron(III)inthecorresponding 3TS1 (IV) sites. Obviously, it is possible for enzyme active sites to direct (12, 13). Hence, the formation of oxyl-ferric species is not the key the actual substrate attack geometry through steric hindrance. factor for the differential reactivity. Thus, the outcome of the enzymatic reaction may depend in a

1232 ∣ www.pnas.org/cgi/doi/10.1073/pnas.1008411108 Ye and Neese Downloaded by guest on September 26, 2021 subtle way on the substrate orientation and this adds another Methods dimension of control to catalysis by metalloenzyme active sites. A truncated model of the ferryl intermediate is employed in the present work In conclusion, the quintet oxo-iron(IV) species is inherently where the iron center is coordinated by two imidazole ligands (representing more reactive toward C–H bond activation than its corresponding His), a monodentate acetate ligand (representing Asp), a bidentate acetate triplet partner. This conclusion is in agreement with previous ligand (representing succinate), and an oxo group. The substrate taurine density functional theory studies (10, 12, 14). Our detailed ana- 2-aminoethane- 1-sulfonic acid) is simply mimicked by ethane. lysis of the electronic structure changes along the reaction coor- We used the hybrid B3LYP density functional (26, 27) in combination with dinate suggests that the real C–H bond cleaving agent is an triple-ζ quality TZVP basis sets (28) on Fe, O, and N as well as SV(P) basis sets oxyl-ferric species that is generated by lengthening of the Fe–oxo (29) on the remaining atoms. The RIJONX approximation (30) was used to bond in the ferryl reactant en route to TS. This enables us to accelerate the calculations in combination with the auxiliary basis sets approach the question of which spin multiplicity of oxo-iron TZV/J (Fe, O, and N) and SV/J (rest) (31, 32). Geometry optimizations were (IV) species has higher reactivity in a completely unique perspec- performed without constraints. The subsequent frequency calculations ver- tive in this work. The main factor contributing to the differential ified that all local minima had only real frequencies and that transition states reactivity is that the two spin states have different requirements (TSs) were distinguished by a single imaginary frequency. The zero-point for the optimal angle at which the substrate should approach the energies, thermal corrections, and entropy terms for the optimized geome- ðFeOÞ2þ core, because distinct electron acceptors are employed tries were obtained from these frequency calculations. Final energy calculations were also performed with the hybrid B3LYP den- on the two surfaces. The H-atom abstraction on the quintet sity functional using the extensively polarized basis sets of triple-ζ quality surface favors the σ-pathway thus requiring an essentially linear including high angular momentum polarization functions (def2-TZVPP) attack. By contrast, a π-channel is operative on the triplet surface (33) for all atoms. The density fitting and chain of spheres (RIJCOSX) approx- that leads to an optimal attack angle near 90°. However, the latter imations (34) have been employed together with the def2-TZVPP/J auxiliary is not possible due to a very large energy penalty if the substrate basis set (35). Following Siegbahn (36), the protein environment was crudely interacts too closely with the supporting ligands of the ferryl modeled by the COSMO model by taking dielectric constant as 4.0. center. This effect is the dominant factor because the energy All calculations were performed with the ORCA program package (37). For – σ required for partially breaking the Fe oxo -bond (quintet sur- an analysis of the changes in the electronic structure of the OH-rebound step, π face) in comparison to the -bond (triplet surface) is very similar. see SI Text. Whereas this is at first glance surprising, it can be explained by the fact that the spin polarization effect stabilizes the HS ferric ACKNOWLEDGMENTS. The authors gratefully acknowledge a grant from the center (formed on the quintet surface) much more strongly than German Science Foundation (NE 690/7-1) and financial support from the CHEMISTRY the LS ferric center that obligatorily evolves on the triplet surface. special research unit SFB 813 (Chemistry at Spin Centers).

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