Inversion of Enantioselectivity of a Mononuclear Non‐Heme Iron(II)‐dependent Hydroxylase by Tuning the Interplay of Metal‐Center Geometry and Protein Structure

Pratter, Sarah; Konstantinovics, Cornelia; Giuro, Cristiana Maria Loredana Di; Leitner, Erich; Kumar, Devesh; Visser, Sam P. de; Grogan, Gideon; Straganz, Grit Daniela

doi:10.1002/ange.201304633

Cited by 25 publications

(18 citation statements)

References 18 publications

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“…The carboxylate group of Asp 145 binds as a monodentate ligand at a distance of 2.01 Å, whereas the succinate carboxylate group binds as a bidentate ligand with distances of 2.20 and 2.30 Å. Again, these distances match previous calculations on similar complexes nicely (Pratter et al, 2013 ). Substrate proline is not bound directly to the iron center but its transferring C–H 4b hydrogen atom is found at a distance of 2.86 Å from the oxo group and hence is positioned in the ideal orientation for oxidation.…”

Section: Resultssupporting

confidence: 85%

How Are Substrate Binding and Catalysis Affected by Mutating Glu127 and Arg161 in Prolyl-4-hydroxylase? A QM/MM and MD Study

Timmins

Visser

2017

Front. Chem.

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Prolyl-4-hydroxylase is a vital enzyme for human physiology involved in the biosynthesis of 4-hydroxyproline, an essential component for collagen formation. The enzyme performs a unique stereo- and regioselective hydroxylation at the C4 position of proline despite the fact that the C5 hydrogen atoms should be thermodynamically easier to abstract. To gain insight into the mechanism and find the origin of this regioselectivity, we have done a quantum mechanics/molecular mechanics (QM/MM) study on wildtype and mutant structures. In a previous study (Timmins et al., 2017) we identified several active site residues critical for substrate binding and positioning. In particular, the Glu127 and Arg161 were shown to form multiple hydrogen bonding and ion-dipole interactions with substrate and could thereby affect the regio- and stereoselectivity of the reaction. In this work, we decided to test that hypothesis and report a QM/MM and molecular dynamics (MD) study on prolyl-4-hydroxylase and several active site mutants where Glu127 or Arg161 are mutated for Asp, Gln, or Lys. Thus, the R161D and R161Q mutants give very high barriers for hydrogen atom abstraction from any proline C–H bond and therefore will be inactive. The R161K mutant, by contrast, sees the regio- and stereoselectivity of the reaction change but still is expected to hydroxylate proline at room temperature. By contrast, the Glu127 mutants E127D and E127Q show possible changes in regioselectivity with the former being more probable to react compared to the latter.

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Section: Resultssupporting

confidence: 85%

How Are Substrate Binding and Catalysis Affected by Mutating Glu127 and Arg161 in Prolyl-4-hydroxylase? A QM/MM and MD Study

Timmins

Visser

2017

Front. Chem.

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“…9 Recently, HMS was successfully engineered to produce R-HMA. 10 X-ray crystal structures for only the HMS-Co(II)-HMA complex (PDB code: 2R5V 7 ), HPPD inhibitor complexes (PDB codes: 1T47, 1SQI, 1TFZ, 1TG5 11, 12 ) and select other ligands (PDB codes: 1CJX, 3ISQ 13 ) have been solved. Thus, the available structural data provide only indirect information on the position of HPP binding within the active sites.…”

Section: Hmsmentioning

confidence: 99%

Role of Substrate Positioning in the Catalytic Reaction of 4-Hydroxyphenylpyruvate Dioxygenase—A QM/MM Study

et al. 2014

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Ring hydroxylation and coupled rearrangement reactions catalyzed by 4-hydroxyphenylpyruvate dioxygenase were studied with the QM/MM method ONIOM(B3LYP:AMBER). For electrophilic attack of the ferryl species on the aromatic ring, five channels were considered: attacks on the three ring atoms closest to the oxo ligand (C1, C2, C6) and insertion of oxygen across two bonds formed by them (C1-C2, C1-C6). For the subsequent migration of the carboxymethyl substituent, two possible directions were tested (C1→C2, C1→C6) and two different mechanisms were sought (step-wise radical, single-step heterolytic). In addition, formation of an epoxide (side)product and benzylic hydroxylation, as catalyzed by the closely related hydroxymandelate synthase, were investigated. From the computed reaction free energy profiles it follows that the most likely mechanism of 4-hydroxyphenylpyruvate dioxygenase involves electrophilic attack on the C1 carbon of the ring and subsequent singlestep heterolytic migration of the substituent. Computed values of the kinetic isotope effect for this step are inverse, consistent with available experimental data. Electronic structure arguments for the preferred mechanism of attack on the ring are also presented.

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“…[20] This change in TS2 barrier leads to fundamental differences in substrate activation and consequently product distributions. However, the subsequent dehydrogenation of hydrogen atom abstraction via TS2 encounters significantly larger barriers of well over DG + E solv > 25 kcal mol À1 , and consequently will be difficult.…”

Section: Angewandte Chemiementioning

confidence: 99%

Comparison of the Reactivity of Nonheme Iron(IV)–Oxo versus Iron(IV)–Imido Complexes: Which is the Better Oxidant?

et al. 2013

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Inversion of Enantioselectivity of a Mononuclear Non‐Heme Iron(II)‐dependent Hydroxylase by Tuning the Interplay of Metal‐Center Geometry and Protein Structure

Abstract: a first step in making enzymatic MNH Fe II centers accessible as a resource for tailor-made biosynthetic oxygenations.

Cited by 25 publications

References 18 publications

How Are Substrate Binding and Catalysis Affected by Mutating Glu127 and Arg161 in Prolyl-4-hydroxylase? A QM/MM and MD Study

How Are Substrate Binding and Catalysis Affected by Mutating Glu127 and Arg161 in Prolyl-4-hydroxylase? A QM/MM and MD Study

Role of Substrate Positioning in the Catalytic Reaction of 4-Hydroxyphenylpyruvate Dioxygenase—A QM/MM Study

Comparison of the Reactivity of Nonheme Iron(IV)–Oxo versus Iron(IV)–Imido Complexes: Which is the Better Oxidant?

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