Are Vanadium Intermediates Suitable Mimics in Non-Heme Iron Enzymes? An Electronic Structure Analysis

Abstract

Vanadyl intermediates are frequently used as mimics for the fleeting Fe(IV)=O intermediate in non-heme iron enzymes that catalyze C–H activation. Using density functional theory and correlated wavefunction theory, we investigate the degree to which vanadium mimic electronic structure is comparable to catalytic iron intermediates. Our calculations reveal crucial structural and energetic differences between vanadyl and ferryl intermediates primarily due to differences in ground spin states of low spin and high spin, respectively. This difference in spin state leads to differences in energetics for accessing isomers that confer activity in non-heme hydroxylase and halogenase enzymes. While interconversion between monodentate and bidentate succinate isomers of the key metal-oxo/hydroxo intermediates is energetically favorable for Fe, it is strongly unfavorable in V mimics. Additionally, isomerization of a terminal metal-oxo between an axial and equatorial position is energetically unfavorable for Fe but favorable for V. Electronic structure analyses to quantify differences in binding strength of Fe and V intermediates to –ketoglutarate and succinate cosubstrates reveal that both cosubstrates bind more strongly to V than Fe. Our work highlights the limits of using low- or intermediate-spin V-based intermediates as mimics in studies of fleeting high-spin Fe intermediates in non-heme iron enzymes.

Optimized Substrate Positioning Enables Switches in C–H Cleavage Site and Reaction Outcome in the Hydroxylation-Epoxidation Sequence Catalyzed by Hyoscyamine 6β-Hydroxylase

“…hydrogens are slightly closer to the metal than the C6 exo hydrogen that is the primary H• donor to the ferryl complex emphasizes the importance of approach angle to HAT efficiency. This notion was first put forth in our work on the halogenase SyrB2, 29 has been supported by multiple computational studies reported since, 32,54,57 and is discussed in more detail below. For the HnH6H•V IV O•succ•7-d1-Hyo complex, the angle relating the g-tensor of the vanadyl spin probe and the metal-deuterium vector was also accessible by analysis of the magnetic field-dependence of the spectra (Figure S15-S21).…”

“…License: CC BY-NC-ND 4.0 configurations of the reactant complexes and by reproducing substrate-association/dissociation dynamics of the ferryl complexes more closely than do the reactant complexes. 5,15,30,36,[51][52][53] More recent computational work employing density function theory (DFT) to estimate relative coordination energies in small active-site models has raised caveats regarding the fidelity of ferryl mimicry by vanadyl, 36,54 but these studies have neither provided alternative explanations for the experimental correlations of structure to reaction dynamics/outcomes nor accounted for the possibility that the larger protein structure could override small energetic differences in intrinsic propensity of the metal and ligands to adopt a certain coordination geometry. We solved structures from crystals of the AbH6H•V IV O•succ•Hyo and AbH6H•V IV O•succ•6-OH-Hyo complexes, the latter the first example of the use of the vanadium cofactor mimic to provide structural insight in a native non-hydroxylation reaction.…”

“…First, it could reflect ambiguous binding of the vanadyl ion in two different orientations in the crystal. 54 However, in refinements to address this possibility, both ligands refined to full occupancy (meaning that both sites are fully occupied at the same time) in both structures, weighing against this explanation. Second, it is possible that a solvent ligand coordinates cis to the vanadyl oxo and becomes chemically equivalent to the oxo as a consequence of photolytic V IV reduction during data collection.…”

“…The larger angle for the preferred hydrogen is consistent with computational studies suggesting that the angle between the Fe IV =O bond and the substrate hydrogen controls the FMOs engaged (s-versus p-pathway) and thus the HAT barrier height, with s-pathway-enabling obtuse angles giving more efficient HAT. 32,54,57 More recent analysis has suggested that, even within the s manifold, the more acute angles lead to slower HAT. To test this idea further, we compared our vanadyl-derived ferryl models of H6H to those that we could generate from reported structures 5,52 (and one other that we solved here 55 ) of other Fe/2OG enzymes.…”

“…This hypothesis comports with the emerging evidence from prior structural analyses that substrate-cofactor disposition is generally crucial to reaction outcome in Fe/2OG oxygenases (Figure 8B). 54 Sterically Perturbing L289F Substitution Disables Epoxidation, Unleashes Second Hydroxylation.…”