The Unexplored Importance of Fleeting Chiral Intermediates in Enzyme-Catalyzed Reactions

Abstract

Decades of extensive research efforts by biochemists, organic chemists, and protein engineers have led to an understanding of the basic mechanisms of essentially all known types of enzymes, but in a formidable number of cases an essential aspect has been overlooked. The occurrence of short-lived chiral intermediates formed by symmetry-breaking of prochiral precursors in enzyme catalyzed reactions has been systematically neglected. We designate these elusive species as fleeting chiral intermediates and analyze such crucial questions as "Do such intermediates occur in homochiral form?" If so, what is the absolute configuration, and why did Nature choose that particular stereoisomeric form, even when the isolable final product may be achiral? Does the absolute configuration of a chiral product depend in any way on the absolute configuration of the fleeting chiral precursor? How does this affect the catalytic proficiency of the enzyme? If these issues continue to be unexplored, then an understanding of the mechanisms of many enzyme types remains incomplete. We have systematized the occurrence of these chiral intermediates according to their structures and enzyme types. This is followed by critical analyses of selected case studies and by final conclusions and perspectives. We hope that the fascinating concept of fleeting chiral intermediates will attract the attention of scientists, thereby opening an exciting new research field.

Enzymatic control over reactive intermediates enables direct oxidation of alkenes to carbonyls by a P450 iron-oxo species

“…S13). 45 The computed QM/MM reaction profile for the P450LA1-catalyzed reaction (Fig. 5) compares well to the DFT modelled enzyme-free truncated model system (Fig.…”

“…Our results demonstrate that confinement 34 is essential to alter and control the intrinsic dynamic behavior of these highly reactive fleeting intermediates formed over the course of these diverging oxidation pathways. The behavior of such intermediates, which indeed can be (pro-)chiral, 45 finally determines the chemoselectivity of the reaction. Mechanistic insights obtained in this work provide useful guidance that may help to further expand this new biocatalytic direct oxidation of alkenes to carbonyl compounds towards more challenging substrates, including internal alkenes 53 or unactivated, aliphatic alkenes.…”

“…4), where the re face of the alkene is exposed to the ironoxo active species (see SI Fig. S13 for calculations involving the alternative minor si-face binding mode, 45 which exhibit significantly higher reaction barriers and thus making it significantly less reactive). These calculations showed that, similarly to the wildtype P450LA1 system, both doublet and quartet radical intermediates aMOx-Int1 can be optimized and characterized as minima on the potential energy surface (Fig.…”

Biocatalytic Baeyer–Villiger Reactions: Uncovering the Source of Regioselectivity at Each Evolutionary Stage of a Mutant with Scrutiny of Fleeting Chiral Intermediates

“…If so, what is the reason for such a switch? These kinds of questions are important for a complete mechanistic understanding of BVMOs and of other enzymes in which chiral fleeting intermediates also occur …”

“…A theoretical analysis based on extensive quantum chemical calculations revealed the underlying mechanism of overriding traditional electronic effects during laboratory evolution on a molecular level at each evolutionary stage. Because the Criegee intermediate is chiral, it belongs to the class of fleeting chiral intermediates, which have remained computationally largely unexplored . The present study includes, for the first time, the calculation of both fleeting chiral species.…”