Stereoelectronic Requirements for Optimal Hydrogen‐Bond‐Catalyzed Enolization

Abstract

Protein crystallographic analysis of the active sites of enolizing enzymes and structural analysis of hydrogen-bonded carbonyl compounds in small molecule crystal structures, complemented by quantum chemical calculations on related model enolization reactions, suggest a new stereoelectronic model that accounts for the observed out-of-plane orientation of hydrogen-bond donors (HBDs) in the oxyanion holes of enolizing enzymes. The computational results reveal that the lone-pair directionality of HBDs characteristic for hydrogen-bonded carbonyls is reduced upon enolization, and the enolate displays almost no directional preference for hydrogen bonding. Positioning the HBDs perpendicular to the carbonyl plane induces strain in the catalyst-substrate complex, which is released upon enolization, resulting in more favorable kinetics and thermodynamics than the in-plane arrangement of HBDs.

The isomerase and hydratase reaction mechanism of the crotonase active site of the multifunctional enzyme (type‐1), as deduced from structures of complexes with 3S‐hydroxy‐acyl‐CoA

“…The perpendicular arrangement of the catalytic water and the OE1(Glu103) with respect to the thioester enolate moiety (Fig. ) is also in agreement with the stereoelectronic rules for this chemistry.…”

“…). The common feature of an active site built on the crotonase fold is the presence of an oxyanion hole, which stabilizes the thioester‐enolate reaction intermediate anion . This important property makes the crotonase fold an attractive framework for the development of new biocatalytic activities, as reported previously .…”

“…). On binding of the thioester oxygen in this oxyanion hole, the p K a of the C2‐proton of the thioester moiety is lowered because the oxyanion hole stabilizes the thioester enolate intermediate . The apo MFE1 active site also contains a sulfate ion bound in the oxyanion hole (Fig.…”

Crotonases: Nature’s Exceedingly Convertible Catalysts

“…The oxyanion is stabilized by conserved hydrogen bonds in the oxyanion hole, and general acid/base catalysis is commonly mediated by aspartate, glutamate, and histidine residues. 76 Although, oxyanion hole-type carbonyl activation is supported by other protein folds, it is possible that the crotonase fold is particularly good at supporting a diverse set of reactions/substrates -this hypothesis could be investigated by systematic designed studies employing both natural and unnatural folds. Many of the crotonase catalyzed reactions described here ( Figure 1B) are analogous to reactions involving carbonyl compounds routinely used in organic synthesis.…”

Human Δ3,Δ2‐enoyl‐CoA isomerase, type 2: a structural enzymology study on the catalytic role of its ACBP domain and helix‐10

“…In ECIs the catalytic residue is a glutamate, which abstracts a proton from the substrate C2 atom, generating a negatively charged enolate . The negative charge of the thioester oxygen atom is stabilized by the OAH . Subsequently, the abstracted proton is transferred to the C4 atom, generating the 2E‐enoyl‐CoA product (Fig.…”