Yongtao Xie

Thiamine diphosphate (ThDP)-dependent enzymes are versatile biocatalysts for carbon–carbon bond formation, yet their catalytic repertoire has been largely confined to linear homologation and intramolecular cyclizations that rely on single-site reactive intermediates such as the Breslow or ketyl radical species. Intermolecular annulation─a powerful strategy for constructing cyclic architectures─has remained elusive within this enzyme class. Here, we report the first intermolecular [3 + 2] annulation catalyzed by a ThDP-dependent enzyme, achieved through a homoenolate-mediated nucleophilic addition pathway. A phylogeny-guided enzyme mining identified a previously uncharacterized enzyme from Variovorax boronicumulans(ThDP-5), and subsequent protein engineering yielded the F484S variant, which exhibits markedly enhanced activity and exquisite stereocontrol. This biocatalytic system exhibits a broad substrate scope (30 examples), accommodating diverse substituted cinnamaldehydes and isatins to afford chiral spirooxindole lactones with excellent stereoselectivity (up to > 20:1 dr and 99% ee). Preparative-scale reactions (2 mmol) validate its synthetic utility. This work expands ThDP-dependent biocatalysis to intermolecular annulations, providing a robust and selective platform for the construction of pharmaceutically relevant spirocyclic architectures.

Chiral organochlorides bearing C-Cl stereocenters are highly desirable motifs in synthetic and medicinal chemistry, yet their direct asymmetric synthesis from simple starting materials via biocatalysis alone remains challenging. Here, we report an integrated organo- and biocatalytic cascade that converts readily available aldehydes into enantiomerically enriched β-chloramines. Under optimized near-neutral conditions, the method delivers a broad array of β-chloramines in high yield (up to 88%) with exceptional enantioselectivity (up to >99:1 enantiomeric ratio). Notably, by simply raising the pH to 9.5, the same cascade system diverges to directly generate chiral aziridines, enabling pH-controlled access to two valuable product classes from a unified platform. The utility of this strategy is further demonstrated by preparative-scale syntheses from inexpensive commercial substrates, followed by divergent linchpin transformations to diverse chiral building blocks, including aziridines, azido amines, and acetoxy amines. Mechanistic studies provide insight into the origins of stereoselectivity and the key factors governing stereochemical outcomes.

Most conventional alkene synthesis reactions (e.g., elimination et al.) inherently favor the formation of thermodynamically more stable E-isomers, posing a long-standing challenge for direct access to Z-alkenes. Here, we report the reprogramming of a thiamine diphosphate (ThDP)-dependent enzyme to catalyze a formal dehalogenative elimination that overrides this intrinsic thermodynamic bias, enabling the direct and selective synthesis of Z-α,β-unsaturated carboxylic acids. In contrast to classical approaches that rely on substrate control, directing groups, or complex ligand architectures, our strategy harnesses the enzyme’s confined active site to achieve kinetic control exclusively via noncovalent interactions─representing a fundamentally distinct and more sustainable approach to stereochemical programming. This transformation diverts the enzyme from its native function in C–C bond formation by channeling the Breslow intermediate toward a homoenolate-mediated pathway, wherein specific noncovalent interactions stabilize the syn-periplanar geometry required for Z-selective dehalogenative elimination. Through rational active-site engineering, the stereochemical trajectory can be inverted to furnish the complementary E-isomer, enabling stereodivergent synthesis from a common scaffold. This work establishes a biocatalytic platform that addresses a critical gap in Z-alkene synthesis, expands the catalytic repertoire of ThDP-dependent enzymes, and provides a sustainable alternative to conventional methodologies.

A green and enantiocomplementary biocatalytic platform for chiral 2,3-dihydroxy aryl ketones has been developed.

Nitrile C–C coupling transformation, currently absent from existing biocatalytic toolbox, is a pivotal objective in enzyme catalysis. Herein, we engineered a ThDP-dependent enzyme to achieve nitrile C–C coupling with in situ-generated acyl anion equivalent, providing a valuable synthetic complement to current enzymatic transformations. This reaction also constitutes the first ThDP-dependent enzymatic addition of aldehyde group to an inert triple bond. Under mildly acidic aqueous conditions, this biocatalytic reaction achieves effective hydrolysis of readily isomerizable imine intermediates. Such a hydrolysis process poses formidable challenges to organic solvent-reliant chemical reactions and has been scarcely explored. In addition, this reaction constitutes an enzyme-catalyzed, oxidant-free strategy for the efficient synthesis of valuable 3-hydroxychromones─key intermediates for accessing flavonol natural products and functional molecules. Related mechanistic studies and preliminary kinetic resolution experiments were also conducted. Our study will encourage further development of biocatalytic systems for nitrile C–C coupling transformations, particularly those challenging to chemical methods.

Traditional biocatalytic cascades typically require discrete enzymes for each synthetic step. Here, we report unprecedented trifunctional imine reductases (IRED) that conduct three sequential transformations—alkene reduction, intramolecular reductive amination, and imine reduction—all within a single catalytic cycle. This elegant single‐enzyme catalytic system directly transforms linear substrates into enantiomerically pure 2‐aryl pyrrolidines via a concerted cascade without intermediate isolation. Combining density functional theory (DFT) calculations and mechanistic studies, we elucidate how the IRED achieves step‐selective catalysis. Our findings establish a proof‐of‐concept for simplifying complex biocatalytic cascades using multifunctional enzymes, offering a powerful strategy to streamline synthetic pathways.

Traditional biocatalytic cascades typically require discrete enzymes for each synthetic step. Here, we report unprecedented trifunctional imine reductases (IRED) that conduct three sequential transformations—alkene reduction, intramolecular reductive amination, and imine reduction—all within a single catalytic cycle. This elegant single-enzyme catalytic system directly transforms linear substrates into enantiomerically pure 2-aryl pyrrolidines via a concerted cascade without intermediate isolation. Combining density functional theory (DFT) calculations and mechanistic studies, we elucidate how the IRED achieves step-selective catalysis. Our findings establish a proof-of-concept for simplifying complex biocascades using multifunctional enzymes, offering a powerful strategy to streamline synthetic pathways.

Taming highly enolizable aldehydes for catalytic asymmetric C-C coupling with nucleophiles remains an elusive challenge compared to widely explored simple alkyl or aryl aldehydes. Herein, we use ThDP-dependent enzymes to realize the direct C-C coupling of highly enolizable 2-phosphonate aldehydes with in situ-generated dynamically reversible nucleophiles (acyl anions). Unlike NHC-mediated reactions that yield complex mixtures of multiple adducts, our enzymatic process selectively produces biologically active β-hydroxy phosphonates with high yields (up to 95%) and excellent enantioselectivities (up to 99% ee). The products can be obtained on gram scales and exhibit rich reactivity for downstream transformations to afford diverse molecules. PfBAL (or its mutant A28G) and PaBAL enzymes serve as enantiocomplementary pairs, enabling the synthesis of both product configurations. Mechanistic studies proved that the entrance directions of the active cavities of these two enzyme pairs were distinct, leading to acyl anions formed from these two enzyme pairs attacking 2-phosphonate aldehydes from different orientations.

Enzymatic catalysis has emerged as an attractive technology for organic synthesis but remains in its infancy for constructing biologically-relevant chiral phosphonates. Here, we disclose a highly chemo- and enantioselective reactions between highly enolizable 2-phosphonate aldehydes and aryl aldehydes for synthesis of chiral phosphonates. This reaction mediated by PfBAL or its mutant A28G features the first example of cross-benzoin reaction involving highly enolizable aldehydes, an elusive challenge previously. Unlike the small molecule NHC-mediated process that give complex mixture of multiple adducts, our enzymatic process selectively gives biologically active hydroxyl ketone-containing phosphonate products in excellent yields and enantioselectivities. The chiral phosphonate products can be obtained on gram scales and carry rich reactivities for further downstream transformation to afford diverse molecules, which will bring more opportunity to the development of novel C-P drugs. This work opened a new avenue in the development of novel cross-benzoin reaction and will stimulate the widespread application of cross-benzoin in organic synthesis.