Mechanistic investigation of repurposed photoenzymes with new-to-nature reactivity

Current Opinion in Green and Sustainable Chemistry, Год журнала: 2025, Номер 52, С. 101009 - 101009

Опубликована: Фев. 27, 2025

Язык: Английский

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Timed batch inputs unlock significantly higher yields for enzymatic cascades

Research Square (Research Square), Год журнала: 2025, Номер unknown

Опубликована: Март 7, 2025

The dynamic properties of enzymatic reaction networks (ERNs) are difficult to predict due the emergence allosteric interactions, product inhibitions and competition for resources, that all only materialize once have been assembled. In batch systems, optimization starting concentrations is challenging, as composition mixture changes continuously, prohibiting optimal conditions full duration reaction. Allowing reagents be added over time would circumvent this limitation, but route has never explored yield complex sequences. Here, we use an active learning workflow construct maximally informative datasets train predictive kinetic models account possible interactions in system. We subsequently show first time, these can used significantly (> 5-fold) improve yields a model cascade (the pentose phosphate pathway) by designing recipe time-dependent inputs each component within network such favors production target metabolite. This work represents generally applicable strategy optimizing networks.

Язык: Английский

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Catalytic Enantioselective Smiles Rearrangement Enabled by the Directed Evolution of P450 Radical Aryl Migratases

Journal of the American Chemical Society, Год журнала: 2025, Номер unknown

Опубликована: Апрель 1, 2025

Despite its synthetic potential, catalytic enantioselective Smiles rearrangement has remained elusive. Through the directed evolution of P450 radical aryl migratases (P450Smiles's), we describe first example rearrangement. A range racemic N-arylsulfonyl-α-chloroamides could be transformed by P450Smiles in an enantioconvergent manner, affording acyclic amide products possessing all-carbon quaternary stereocenter with excellent chemo- and enantioselectivity. Both electron-rich electron-deficient substituents were compatible migrating group, demonstrating this P450-catalyzed is insensitive to electronic properties group. Importantly, our evolved variants capable overriding innate cyclization activity N-alkyl amidyl intermediate, allowing chemoselective reductive formation products. Classical molecular dynamics (MD) simulations revealed unusual enzyme-controlled chemoselectivity stems from restricted conformation within enzyme active site, disfavoring pathway. This new-to-nature biocatalytic asymmetric showcases potential enzymatic enantioselectivity control over highly reactive intermediates eluding small-molecule catalysts.

Язык: Английский

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How to Tell an N from an O: Controlling the Chemoselectivity of Methyltransferases

ACS Catalysis, Год журнала: 2025, Номер unknown, С. 6410 - 6425

Опубликована: Апрель 4, 2025

S-Adenosyl-l-methionine (SAM)-dependent methyltransferases (MTs) are important enzymes in numerous biological pathways. They share a common S N 2 mechanism but act on different nucleophilic substrates vivo. Therefore, MTs have specific chemoselectivity to transfer CH3 onto the correct atom type and substrate. Caffeate O-MT from Prunus persica (PpCaOMT) anthranilate N-MT Ruta graveolens (RgANMT) high similarity regarding their amino acid sequence (>74%). Nevertheless, physiological (caffeate vs anthranilate) attacking nucleophiles (hydroxyl group) strikingly different. We demonstrate that differing is governed by conformational states of two enzymes. O-Methylation catalyzed CaOMTs requires "closed" conformation, whereas ANMTs perform N-methylation an "open" state. rationally designed seven variants for both PpCaOMT RgANMT, which changed original nucleophile preference extents, up full inversion. Interestingly, generated O-selective ANMT variant catalyzes O-methylation considerably faster than wildtype CaOMT. Molecular dynamics (MD) simulations hydrogen/deuterium exchange mass spectrometry (HDX-MS) experiments showed mutations induced changes enzyme modulating open/closed transitions impact corresponding chemoselectivity. Our data show selectivity methyl reaction not solely key residues directly involved rather synergistically modulated conditions.

Язык: Английский

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Unravelling the Potential of Crude Enzyme Extracts for Biocatalyst Entrapment in Metal–Organic Frameworks

ACS Nano, Год журнала: 2025, Номер unknown

Опубликована: Апрель 11, 2025

To bolster the applicability of enzymes as catalysts, it is imperative not only to address their inherent fragility, particularly when used under harsh organic-synthetic reaction conditions, but also mitigate deactivation during purification and enable in a broad range transformations. Currently, process crude enzyme extracts subsequent heterogenization obtain immobilized biocatalysts often leads partial represents, at least part, resource-intensive that driving up overall production efforts. tackle both fragility immobilization, we propose direct use obtained from cell lysis instead pure entrapment metal-organic framework (MOF) structures. We focus on three types with varying sensitivities: aldoxime dehydratase, imine reductase, lipase. evaluate effects different metal sources (Al, Fe, Co, Ni, Cu, Zn), oxidation state counterions, MOF synthesis parameters stability activity Based this, optimize protocols for Fe-MIL-88A, Fe-MIL-100, Zn-MOF-74, Zn-ZIF-8 develop fast-aqueous room temperature Al-MIL-53. Investigation biocatalytic performance enzyme@MOF biocomposites suggests MOFs using can effectively maintain various catalytic reactions, offering perspective an efficient pathway industrial applications.

Язык: Английский

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Sucrose conversion to 5-hydroxymethylfurfural over commercial ion-exchange resin

Biomass and Bioenergy, Год журнала: 2025, Номер 199, С. 107940 - 107940

Опубликована: Апрель 28, 2025

Язык: Английский

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Directed Evolution Enables Dynamic Control of Transient Intermediates for Anti-Markovnikov Wacker-Tsuji-Type Oxidation of Aliphatic Alkenes

Опубликована: Июль 29, 2024

Controlling the dynamic behavior of highly reactive fleeting intermediates is an intriguing concept to generate selectivity in competing reaction pathways. Here we report directed evolution a P450 enzyme for anti-Markovnikov-selective Wacker-Tsuji-type oxidation aliphatic alkenes. This long-standing challenge catalysis due its unfavorable energetics and reactions. The evolved aMOx-A performs anti-Markovnikov alkenes with several hundred turnovers possesses kinetic parameters comparable those average natural enzymes. biocatalyst guides oxo-transfer process through multiple reactions, including bifurcating pathways that originate after rate-limiting transition state. Chemoselectivity arises from controlling substrate conformations as well dynamics intermediates. includes accessing carbocation intermediate does not correspond minimum on potential energy surface. Engineering highlights how optimizes unique catalytic processes have largely eluded efficient catalysis.

Язык: Английский

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Active learning maps the emergent dynamics of enzymatic reaction networks.

Опубликована: Авг. 29, 2024

The dynamic properties of enzymatic reaction networks (ERNs) are difficult to predict due the emergence allosteric interactions, product inhibitions and competition for resources, that all only materialize once have been assembled. Combining experimental kinetics studies with computational modelling allows us extract information on these emergent build predictive models. Here, we utilized pentose phosphate pathway demonstrate previously reported approaches construct maximally informative datasets can be significantly improved by pulsing both enzymes substrates into microfluidic flow reactors (instead only). Our method augments available from online databases, map behaviours a network.

Язык: Английский

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Recent progress in asymmetric radical reactions enabled by chiral iron catalysts

Chemical Communications, Год журнала: 2024, Номер unknown

Опубликована: Янв. 1, 2024

In this highlight, we discuss recently developed chiral iron catalysts and their application in radical asymmetric reactions for the construction of axial or central chirality.

Язык: Английский

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