Elucidation of factors shaping reactivity of 5'-deoxyadenosyl - a prominent organic radical in biology.

This study investigates the factors modulating the reactivity of 5'-deoxyadenosyl (5'dAdo˙) radical, a potent hydrogen atom abstractor that forms in the active sites of radical SAM enzymes and that otherwise undergoes a rapid self-decay in aqueous solution. Here, we compare hydrogen atom abstraction (HAA) reactions between native substrates of radical SAM enzymes and 5'dAdo˙ in aqueous solution and in two enzymatic microenvironments. With that we reveal that HAA efficiency of 5'dAdo˙ is due to (i) the formation of 5'dAdo˙ in a pre-ordered complex with a substrate, which attenuates the unfavorable effect of substrate:5'dAdo˙ complex formation, and (ii) the prevention of the conformational changes associated with self-decay by a tight active-site cavity.

Radical ligand transfer: mechanism and reactivity governed by three-component thermodynamics.

Here, we demonstrate that the relationship between reactivity and thermodynamics in radical ligand transfer chemistry can be understood if this chemistry is dissected as concerted ion-electron transfer (cIET). Namely, we investigate radical ligand transfer reactions from the perspective of thermodynamic contributions to the reaction barrier: the diagonal effect of the free energy of the reaction, and the off-diagonal effect resulting from asynchronicity and frustration, which we originally derived from the thermodynamic cycle for concerted proton-electron transfer (cPET). This study on the OH transfer reaction shows that the three-component thermodynamic model goes beyond cPET chemistry, successfully capturing the changes in radical ligand transfer reactivity in a series of model Fe-OH⋯(diflouro)cyclohexadienyl systems.

Properties of the Reactants and Their Interactions within and with the Enzyme Binding Cavity Determine Reaction Selectivities. The Case of Fe(II)/2-Oxoglutarate Dependent Enzymes.

Fe(II)/2-oxoglutarate dependent dioxygenases (ODDs) share a double stranded beta helix (DSBH) fold and utilise a common reactive intermediate, ferryl species, to catalyse oxidative transformations of substrates. Despite the structural similarities, ODDs accept a variety of substrates and facilitate a wide range of reactions, that is hydroxylations, desaturations, (oxa)cyclisations and ring rearrangements. In this review we present and discuss the factors contributing to the observed (regio)selectivities of ODDs.

Enzyme Multifunctionality by Control of Substrate Positioning Within the Catalytic Cycle-A QM/MM Study of Clavaminic Acid Synthase.

Clavaminic acid synthase from Streptomyces clavuligerus is an Fe /2-oxoglutarate-dependent dioxygenase, crucial for the biosynthesis of the β-lactamase inhibitor clavulanic acid. It catalyses three consecutive oxidative reactions, that is, hydroxylation, cyclisation and desaturation, in a single binding cavity. As follows from the results of this QM/MM study, CAS versatility and selectivity depends on the binding cavity, which interplays differently with the substrate for each reaction.

It takes two to tango - The case of thebaine 6-O-demethylase.

Thebaine 6-O-demethylase (T6ODM) is an Fe(II)/2-oxoglutarate-dependent dioxygenase catalysing two oxidative O-demethylation reactions in morphine biosynthesis. Its crystal structure revealed a large active site pocket which is at least two times larger than necessary to accommodate a substrate (thebaine or oripavine) molecule. Since so far no crystal structures have been obtained for enzyme-substrate complex, which is necessary to explain the enzyme regiospecificity towards the C6-bound methoxy group, in this work we used computational methods and multi-parametric surface plasmon resonance measurements to elucidate the most likely structure of this complex and the reaction mechanism starting therefrom.

Regioselectivity of hyoscyamine 6β-hydroxylase-catalysed hydroxylation as revealed by high-resolution structural information and QM/MM calculations.

Hyoscyamine 6β-hydroxylase (H6H) is a bifunctional non-heme 2-oxoglutarate/Fe2+-dependent dioxygenase that catalyzes the two final steps in the biosynthesis of scopolamine. Based on high resolution crystal structures of H6H from Datura metel, detailed information on substrate binding was obtained that provided insights into the onset of the enzymatic process. In particular, the role of two prominent residues was revealed - Glu-116 that interacts with the tertiary amine located on the hyoscyamine tropane moiety and Tyr-326 that forms CH-π hydrogen bonds with the hyoscyamine phenyl ring.

On how the binding cavity of AsqJ dioxygenase controls the desaturation reaction regioselectivity: a QM/MM study.

The Fe(II)/2-oxoglutarate-dependent dioxygenase AsqJ from Aspergillus nidulans catalyses two pivotal steps in the synthesis of quinolone antibiotic 4'-methoxyviridicatin, i.e., desaturation and epoxidation of a benzodiazepinedione.

Crystal structure of thebaine 6-O-demethylase from the morphine biosynthesis pathway.

Thebaine 6-O-demethylase (T6ODM) from Papaver somniferum (opium poppy), which belongs to the non-heme 2-oxoglutarate/Fe(II)-dependent dioxygenases (ODD) family, is a key enzyme in the morphine biosynthesis pathway. Initially, T6ODM was characterized as an enzyme catalyzing O-demethylation of thebaine to neopinone and oripavine to morphinone. However, the substrate range of T6ODM was recently expanded to a number of various benzylisoquinoline alkaloids.

DFT study of the mechanism of manganese quercetin 2,3-dioxygenase: quest for origins of enzyme unique nitroxygenase activity and regioselectivity.

Quercetin 2,3-dioxygenase (QDO) is an enzyme which accepts various transition metal ions as cofactors, and cleaves the heterocyclic ring of quercetin with consumption of dioxygen and release of carbon monoxide. QDO from B. subtilis that binds Mn(II) displays an unprecedented nitroxygenase activity, whereby nitroxyl (HNO) is incorporated into quercetin cleavage products instead of dioxygen.