O2 Activation by Non-Heme Iron Enzymes

Abstract

The non-heme Fe enzymes are ubiquitous in nature and perform a wide range of functions involving O₂ activation. These had been difficult to study relative to heme enzymes; however, spectroscopic methods that provide significant insight into the correlation of structure with function have now been developed. This Current Topics article summarizes both the molecular mechanism these enzymes use to control O₂ activation in the presence of cosubstrates and the oxygen intermediates these reactions generate. Three types of O₂ activation are observed. First, non-heme reactivity is shown to be different from heme chemistry where a low-spin Fe^III-OOH non-heme intermediate directly reacts with substrate. Also, two subclasses of non-heme Fe enzymes generate high-spin Fe^IV═O intermediates that provide both σ and π frontier molecular orbitals that can control selectivity. Finally, for several subclasses of non-heme Fe enzymes, binding of the substrate to the Fe^II site leads to the one-electron reductive activation of O₂ to an Fe^III-superoxide capable of H atom abstraction and electrophilic attack.

Figure 1

Panel A: Representative low temperature MCD spectra for (from L to R) 6C octahedral, 5C square pyramidal, 5C trigonal bipyramidal, and 4C distorted tetrahedral non-heme Fe (NHFe) complexes. Panel B: MCD spectra for resting PAH (black) overlaid with spectra for (from L to R) PAH-L-Phe (blue), PAH-pterin (green), and PAH-L-Phe-pterin (red). Only coordination of both cosubstrates leads to a 5C site. Adapted from ref .

Figure 2

The general mechanistic strategy for NHFe enzymes, where a coordination position for O₂ is only available when all cosubstrates are bound to the active site. Adapted from ref .

Figure 3

Schematic showing the H-bonding and steric contributions to water loss and 5C site formation in FIH upon substrate (C- terminal transactivation domain, CAD) binding. Adapted from ref .

Figure 4

Schematic comparison of facial triad coordination for the classes of NHFe enzymes that have a second-sphere residue hydrogen bonding to the coordinated carboxylate (αKG dependent enzymes, shown at left, and the extradiol dioxygenases), leading to monodentate coordination, and those without a hydrogen bonding residue (pterin dependent enzymes, shown at right, and the Rieske dioxygenases), which leads to bidentate carboxylate coordination.

Figure 5

A: NRVS spectra (top) and simulations (bottom) for low-spin ferric BLM showing an axial hydroxide bound to the Fe. B: NRVS data (top) and simulations (bottom) for ABLM. The three-peak pattern in the data can only be modeled by a low-spin Fe^III-OOH structure. C: Schematic depictions of vibrations assigned in B. Adapted from ref .

Figure 6

A: Transition state for the direct H-atom abstraction of a proton by ABLM, which is late in O-O cleavage and early in C-H abstraction. B: The hydroperoxo σ* FMO of the low-spin ABLM reactant (left), which polarizes into a hydroxyl radical and Fe^IV=O at the transition state (right). Adapted from ref .

Figure 7

Top: NRVS spectra for SyrB2-Cl (green) and SyrB2-Br (red). Middle: DFT NRVS simulations derived from the structure that best reproduces the experimental data. Bottom: Experimentally derived structure of the Fe^IV=O intermediate of SyrB2, which has a 5C trigonal bipyramidal geometry and an ~C₃ axis. Adapted from ref .

Figure 8

A: Absorption (top) and MCD data (bottom) for [Fe^IV(O)(TMG₃tren)]²⁺. B: LF and CT transitions of [Fe^IV(O)(TMG₃tren)]²⁺ C: FMOs of [Fe^IV(O)(TMG₃tren)]²⁺ at the transition state demonstrating Fe^III-O^•− character and the different channels for reactivity. Adapted from ref .

Figure 9

FMOs of the Fe^IV=O intermediate in HPPD (left) and HmaS (right), demonstrating that different substrate orientations lead to different reactivities using different FMOs. Adapted from ref .

Figure 10

Reaction coordinate for O₂ activation of SyrB2, leading to a 5C trigonal bipyramidal Fe^IV=O intermediate (as in Figure 7) with an Fe-O bond perpendicular to the substrate C-H bond. The contour gives the π* FMO used for reactivity. Adapted from ref .

Figure 11

Comparison of the free energies for the one-electron reduction of NO/O₂ by Fe^II (middle) and for the formation of a Fe^III-NO/O₂ complex (right) for a resting facial triad and for substrate-bound IPNS. Sulfur donation stabilizes formation of an Fe^III-O₂⁻ complex by 25.9 kcal/mol relative to a resting facial triad. Adapted from ref .

Figure 12

FMO of Fe^III-ACV-O₂. Inset: Schematic of the unoccupied FMO, showing its favorable orientation for H-atom abstraction from the substrate β methylene carbon. Adapted from ref .

Scheme 1

Kinetic model including the general mechanistic strategy in the αKG dependent dioxygenases

Scheme 2

Possible electronic structures for the activated Fe-O₂ species in the extradiol dioxygenases.