Braca

In this study, we capitalize on the versatility offered by genetic code expansion to develop a proficient photoenzyme for [2 + 2]-cycloadditions that is enantiocomplementary to our recently developed photoenzyme EnT1.3. By repositioning the genetically programmed benzophenone photosensitizer and subsequent directed evolution, we have developed an efficient, highly selective and oxygen tolerant enzyme for a range of [2 + 2]-cycloadditions.

Abstract

The combination of genetic code expansion and directed evolution has recently given rise to enantioselective photoenzymes for [2 + 2]-cycloadditions of quinolone and indole derivatives. However, the enzymes reported to date only allow access to one enantiomeric series of the strained cyclobutane products. Here, guided by a crystal structure of our previously engineered enzyme EnT1.3, we show how judicious repositioning of the genetically programmed benzophenone photosensitizer affords an enantiocomplementary [2 + 2]-cyclase, CEnT1.0. Following directed evolution, a proficient and oxygen-tolerant photoenzyme (CEnT1.4) emerged that promotes [2 + 2]-cycloadditions of a quinolone derivative with exquisite enantiocontrol (99% e.e.) and substantially enhanced regioselectivity compared with EnT1.3 (r.r. 62:1 vs. 9:1). Structural analysis of CEnT1.4, coupled with molecular dynamic simulations, reveals a well-sculpted active site pocket that pre-organises the substrate for regio- and enantioselective catalysis. This study highlights the versatility offered by genetically programmed (photo)catalytic elements when developing enzymes for altered stereochemical outcomes.

This study introduces a class of artificial metalloenzymes (ArMs) featuring octahedral iron complexes. Two such cofactors are prepared and embedded in protein scaffolds via biotin—streptavidin technology to create artificial metalloenzymes for enantioselective 1,3-nitrogen migration. The resulting ArMs showed mutant-dependent activity, highlighting synergy between the metal center and protein environment.

Abstract

Octahedral complexes with linear tetradentate ligands are effective catalysts for various chemical transformations, with the cis-α geometry typically being the active form. These complexes possess stereogenic metal centers, adopting either Λ- or Δ-configurations. We hypothesized that embedding these complexes within the chiral environment of protein scaffolds could yield artificial metalloenzymes (ArMs) with both an enantioenriched metal center and a chiral catalytic pocket. This combination could potentially act synergistically, enhancing catalytic reactivity and selectivity, thereby producing more powerful and tunable catalysts compared to the complexes alone. In this study, we designed two cofactors featuring octahedral Fe complexes: cofactor 1, which, due to its rigid chiral backbone, exclusively forms the Δ-configuration, and cofactor 2, which forms a racemic mixture. Using biotin-streptavidin technology, we developed ArMs based on these cofactors and evaluated them in a 1,3-nitrogen migration reaction to produce chiral α-amino acids. Both ArMs exhibited protein mutant-dependent reactivity and selectivity, with cofactor 1 demonstrating notable synergistic or conflicting effects between the protein scaffold and the metal-centered configuration. This work highlights the potential of developing ArMs with stereogenic metal centers for highly selective catalysis.

Most practical fluorination reagents deliver a fluorine atom either as a nucleophile (F–) or as an electrophile (F+). In contrast, bench-stable radical fluorine (F•) reagents are relatively less common and the vast majority of ‘radical fluorinations’ involve reactions of carbon-centered radicals with electrophilic fluorination reagents. Here, we disclose that silver (II) fluoride (AgF2) in acetonitrile is a mild source of F• that can be leveraged for the synthesis of a variety of high-value organofluorine compounds from abundantly available reactants such as alkanes, alkenes, and carboxylic acids, as well as from pharmaceutically relevant heterocycles such as indoles and benzofurans. This platform technology obviates the need for expensive catalysts and fluorinating reagents that are typically necessary to accomplish these transformations and relies on the use of AgF2 in acetonitrile as the sole reagent under mild conditions.

This review evaluates engineered (semi)autonomous cell systems for the biosynthesis and incorporation of noncanonical amino acids (ncAAs) into proteins. While semi-autonomous cells convert supplied precursors into ncAAs autonomous cells integrate biosynthetic pathways that produce these building blocks intracellularly. Such integrated approaches significantly reduce process costs, can increase protein yields, and overcome challenges such as the limited membrane permeability of ncAAs.

Autonomous cells are engineered biological systems capable of biosynthesising and directly incorporating noncanonical amino acids (ncAAs) into proteins. These systems have the potential to extend the applicability of the genetic code to enable large-scale fermentative production of proteins carrying ncAAs. This work evaluates approaches for the generation of autonomous and semi-autonomous cells. Semi-autonomous cells rely on the external addition of a precursor, which is enzymatically converted in vivo to an ncAA that is directly incorporated. In contrast, autonomous cells have a metabolic system that produces and directly incorporates an ncAA in vivo. Through a critical evaluation of the state of the art, the reader is provided with an opinion on the future development of the field.

Visible light photocatalysis that exploits the reactivity of molecules at their excited state has induced a paradigm shift in organic synthesis by enabling unique chemical transformations, but controlling their enantioselectivity has proven difficult. A promising strategy involves linking a synthetic transition metal photocatalyst within the chiral architecture of a biomolecule to create a highly selective artificial photoenzyme. However, such a biohybrid system that combines the merits of biocatalysis and metallo-photocatalysis to promote abiological reactions fueled by visible light with high enantioselectivity is still unknown. Here, we report on an artificial metallo-photoDNAzyme resulting from covalently anchoring a blue light absorbing iridium-based photocatalyst within a double-stranded DNA helix that exhibits efficient triplet-triplet energy transfer and high levels of enantioselectivity in [2+2] intramolecular cycloadditions.

Nature Catalysis, Published online: 24 June 2025; doi:10.1038/s41929-025-01347-0

Remote C–H bond formation via photoenzymatic hydrogen atom transfer has enabled the precise installation of remote stereocentres but is still in its infancy. Here, the authors report the photoenzymatic stereoablative enantioconvergence of γ-chiral oximes using repurposed flavin-dependent ene-reductases.

The integration between visible-light photocatalysis and biocatalysis has the potential to greatly improve modern organic synthesis by combining the production of valuable synthetic intermediates under mild photochemical conditions with the high degree of stereoselectivity reached by enzymes. The review collects the recent efforts in the photoenzymatic asymmetric synthesis of alcohols and amines.

The stereocontrolled formation of CO and CN bonds is important for the preparation of agrochemicals, pharmaceuticals, fragrances, and flavors. In this field, enzymes have become a valid alternative to transition metal complexes and organocatalysts due to their high degree of stereoselectivity and desirable sustainability features. In this scenario, the increasing interest toward sustainable and efficient chemical processes has also led to the combination of complementary catalytic systems. The merge of biocatalysis and photocatalysis is among the most recent approaches. While photocatalysis allows the production of valuable synthetic intermediates and reactive species under mild conditions, biocatalysis exploits highly specific enzymes to catalyze reactions with high (stereo)-selectivity and minimal byproduct formation. This review summarizes the progress in photoenzymatic catalysis (also photobiocatalysis), emphasizing their complementary mechanisms in producing chiral alcohols and amines, and highlights how such integration not only enhances the sustainability of catalytic systems but also expands the scope of reactions accessible under milder and ultimately more sustainable conditions.

Nature, Published online: 18 June 2025; doi:10.1038/s41586-025-09136-2

We present a computational approach to the design of high-efficiency enzymes with catalytic parameters comparable to natural enzymes, enabling programming of stable, high-efficiency, new-to-nature Kemp elimination enzymes through minimal experimental effort.

The protein LmrR is a versatile scaffold to design artificial metalloenzymes. Here, we show that the M8D/A92E (DE) mutations that result in a much more efficient catalyst, destabilize the protein dimerization interface and alter the conformation landscape when binding metal cofactor and substrates are shown. These results highlight the intricate relations between protein, metal cofactor, and substrates in defining catalytic efficiency.

Artificial metalloenzymes (ArM) hold great potential for the sustainable catalysis of complex new-to-nature reactions. To efficiently improve the catalytic efficacy of ArMs, a rational approach is desirable, requiring detailed molecular insight into their conformational landscape. Lactococcal multidrug resistance regulator (LmrR) is a multipurpose ArM scaffold protein that, when bound to the Cu(II)-phenanthroline cofactor, catalyzes the Friedel–Crafts alkylation (FCA) of indoles. Previously, the M8D and A92E mutations are found to increase the efficiency of this reaction, but a molecular explanation has been lacking. The impact of these two activating mutations on the conformational landscape of LmrR in its apo, cofactor- and substrate-bound states is determined. The mutations cause a marked destabilization of the dimerization interface, resulting in a more open central hydrophobic cavity and a dynamic equilibrium between dimer and monomer LmrR is found. While mutant and wild-type have similar pocket conformation in the cofactor-bound state, the mutant shows a distinct interaction with the substrate. Our results suggest that increased retention of the catalytic cofactor and widened plasticity improve the activity of the mutant. Ultimately, these results help elucidating the intricate relationships between conformational dynamics of the protein scaffold, cofactor, and substrates that define catalytic activity.

Visible light photocatalysis that exploits the reactivity of molecules at their excited state has induced a paradigm shift in organic synthesis by enabling unique chemical transformations, but controlling their enantioselectivity has proven difficult. A promising strategy involves linking a synthetic transition metal photocatalyst within the chiral architecture of a biomolecule to create a highly selective artificial photoenzyme. However, such a biohybrid system that combines the merits of biocatalysis and metallo-photocatalysis to promote abiological reactions fueled by visible light with high enantioselectivity is still unknown. Here, we report on an artificial metallo-photoDNAzyme resulting from covalently anchoring a blue light absorbing iridium-based photocatalyst within a double-stranded DNA helix that exhibits efficient triplet-triplet energy transfer and high levels of enantioselectivity in [2+2] intramolecular cycloadditions.

Nature Chemistry, Published online: 04 June 2025; doi:10.1038/s41557-025-01842-8

Proteins with small structural modifications at specific sites are valuable, yet challenging to access by chemical methods. Now, tyrosine-selective single-atom modifications on proteins have been achieved by C–H functionalization using a rationally designed selenoxide to introduce a versatile selenonium linchpin for further transformations.

Nature Chemistry, Published online: 29 May 2025; doi:10.1038/s41557-025-01835-7

Gold redox catalysis is an attractive synthetic method but challenging due to the high redox potential of Au(I)/Au(III). Now, a bidentate N-ligand-assisted gold redox catalysis using H2O2 as oxidant has been developed. It can be applied to various coupling reactions, including C(sp)–C(sp) cross-coupling, alkynylative cyclization and bicyclization coupling.

Nature, Published online: 28 May 2025; doi:10.1038/s41586-025-09178-6

Electricity-driven enzymatic dynamic kinetic oxidation

By harnessing the synergy between enzymes and photoredox catalysts, cooperative photobiocatalysis has recently emerged as a promising strategy for developing stereoselective radical reactions. While various cofactor-dependent enzymes have been repurposed, the use of cofactor-independent enzymes in such cooperative catalysis without requiring expensive cofactors remains rare. Herein, we report the successful repurposing of class I aldolases, a prominent family of naturally occurring, cofactor-independent enzymes, to catalyze unnatural radical α-alkylation of aldehydes in a highly enantioselective fashion. Through directed evolution of Escherichia coli 2-deoxy-D-ribose-5-phosphate aldolase (EcDERA), we developed an effective radical alkylase bearing five mutations and inverted π–facial selectivity relative to wild-type EcDERA, allowing a range of aldehydes to couple with α-iodoesters, α-iodoketones and α-iodonitriles with excellent enantiocontrol. This study represents the first demonstration of leveraging the nucleophilic enamine intermediate in class I aldolases for radical-mediated stereoselective C–C bond formation. Mechanistic investigations suggested that when irradiated at 440 nm, cooperative catalysis with an exogenous Ir photocatalyst more effectively induces enzymatic enamine radical activity than charge-transfer complex photochemistry. Together, these findings underscore the potential of class I aldolases to enable general and stereoselective new-to-nature radical transformations.

Nature Catalysis, Published online: 16 May 2025; doi:10.1038/s41929-025-01340-7

Transformations from carbenes to olefins have generally been realized with transition metal-catalysed enantioselective methods or artificial metalloenzymes. Here the authors apply asymmetric counteranion-directed photoredox organocatalysis for the highly enantioselective cyclopropanation of styrenes and aliphatic dienes.

This mini-review highlights the use of ene-reductases under visible light to trigger single-electron-reduction-initiated radical reactions, enabling controlled CC, CN, CO, and CS bond formations. These reactions provide significant assistance in addressing long-standing challenges in chemical synthesis.

Flavin-dependent ene-reductases (EREDs) have emerged as powerful biocatalysts for the asymmetric reduction of various substrates. This review focuses on the recent advances in light-induced electron transfer and subsequent reduction reactions mediated by EREDs. Upon photoexcitation, the flavin cofactor transitions to an excited state, significantly enhancing its reduction potential. Mechanistic insights into how light activation alters the redox properties of EREDs are discussed, leading to more efficient catalysis. The review also highlights the broadened application scope of photoexcited EREDs in organic synthesis. Additionally, the challenges and future directions in optimizing these light-driven biocatalytic processes are explored. This overview provides a foundation for developing novel, light-controlled enzymatic systems with enhanced catalytic performance.

Through the directed evolution of an underexploited nonheme Fe extradiol dioxygenase, we developed a unified cooperative photobiocatalytic strategy to allow for three types of enantioconvergent radical transformations, including azidation, thiocyanation, and isocyanation. Computational studies based on density functional theory (DFT) and molecular dynamics (MD) simulations suggested a π-facial selective radical rebound mechanism as the enantiodeter

Abstract

Cooperative catalysis with an enzyme and a small-molecule photocatalyst has recently emerged as a potentially general activation mode to advance novel biocatalytic reactions with synthetic utility. Herein, we report cooperative photobiocatalysis involving an engineered nonheme Fe enzyme and a tailored photoredox catalyst to achieve enantioconvergent decarboxylative azidation, thiocyanation, and isocyanation of redox-active esters via a radical mechanism. We repurposed and further evolved metapyrocatechase (MPC), a nonheme Fe extradiol dioxygenase not previously studied in new-to-nature biocatalysis, for the enantioselective C─N₃, C─SCN, and C─NCO bond formation via an enzymatic Fe─X intermediate (X═N₃, NCS, and NCO). A range of primary, secondary, and tertiary alkyl radical precursors were effectively converted by our engineered MPC, allowing the syntheses of organic azides, thiocyanates, and isocyanates with good to excellent enantiocontrol. Further derivatization of these products furnished valuable compounds including enantioenriched amines, triazoles, ureas, and SCF₃-containing products. DFT and MD simulations shed light on the mechanism as well as the binding poses of the alkyl radical intermediate in the enzyme active site and the π-facial selectivity in the enantiodetermining radical rebound. Overall, cooperative photometallobiocatalysis with nonheme Fe enzymes provides a means to develop challenging asymmetric radical transformations eluding small-molecule catalysis.

Halogen-atom transfer (XAT) is an effective method for generating carbon radicals from alkyl halides. While stannanes and silanes have shown potential, their toxicity and secondary reactivity necessitate the search for better alternatives. Recently, carbon-based α amino alkyl radicals have emerged as possible substitutes; however, their generation under photochemical conditions requires the use of photocatalysts. In this work, we present a novel XAT method that eliminates the need for photocatalysts by utilizing the direct excitation of a monofluoroacetoxy ligand-containing hypervalent iodine(III) reagent. This approach allows nucleophilic monofluoromethyl radicals to efficiently form carbon radicals from activated alkyl halides. These radicals can subsequently functionalize unactivated alkenes through atom transfer radical addition (ATRA). Our mechanistic studies provide insights into the generation of monofluoromethyl radical and its unique reactivity. This work highlights the untapped potential of simple, single carbon atom-containing radicals for effective alkyl radical generation.