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The interconversion of monoterpenes is facilitated by a complex network of carbocation rearrangement pathways. Controlling these isomerization pathways is challenging when using common Brønsted and Lewis acid catalysts, which often produce product mixtures that are difficult to separate. In contrast, natural monoterpene cyclases exhibit high control over the carbocation rearrangement reactions but are reliant on phosphorylated substrates. In this study, we present engineered squalene‐hopene cyclases from Alicyclobacillus acidocaldarius (AacSHC) that catalyze the challenging isomerization of monoterpenes with unprecedented precision. Starting from a promiscuous isomerization of (+)‐β‐pinene, we first demonstrate noticeable shifts in the product distribution solely by introducing single point mutations. Furthermore, we showcase the tuneable cation steering by enhancing (+)‐borneol selectivity from 1 % to >90 % (>99 % de) aided by iterative saturation mutagenesis. Our combined experimental and computational data suggest that the reorganization of key aromatic residues leads to the restructuring of the water network that facilitates the selective termination of the secondary isobornyl cation. This work expands our mechanistic understanding of carbocation rearrangements and sets the stage for target‐oriented skeletal reorganization of broadly abundant terpenes. [ABSTRACT FROM AUTHOR]

Terpenoids are built from isoprene building blocks and have numerous biological functions. Selective late‐stage modification of their carbon scaffold has the potential to optimize or transform their biological activities. However, the synthesis of terpenoids with a non‐natural carbon scaffold is often a challenging endeavor because of the complexity of these molecules. Herein we report the identification and engineering of (S)‐adenosyl‐l‐methionine‐dependent sterol methyltransferases for selective C‐methylation of linear terpenoids. The engineered enzyme catalyzes selective methylation of unactivated alkenes in mono‐, sesqui‐ and diterpenoids to produce C11, C16 and C21 derivatives. Preparative conversion and product isolation reveals that this biocatalyst performs C−C bond formation with high chemo‐ and regioselectivity. The alkene methylation most likely proceeds via a carbocation intermediate and regioselective deprotonation. This method opens new avenues for modifying the carbon scaffold of alkenes in general and terpenoids in particular. [ABSTRACT FROM AUTHOR]

Copyright of Angewandte Chemie is the property of Wiley-Blackwell and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Enantioselective synthesis of chiral alcohols through asymmetric addition of water across an unactivated alkene is a highly sought‐after transformation and a big challenge in catalysis. Herein we report the identification and directed evolution of a fatty acid hydratase from Marinitoga hydrogenitolerans for the highly enantioselective hydration of styrenes to yield chiral 1‐arylethanols. While directed evolution for styrene hydration was performed in the presence of heptanoic acid to mimic fatty acid binding, the engineered enzyme displayed remarkable asymmetric styrene hydration activity in the absence of the small molecule activator. The evolved styrene hydratase provided access to chiral alcohols from simple alkenes and water with high enantioselectivity (>99 : 1 e.r.) and could be applied on a preparative scale. [ABSTRACT FROM AUTHOR]

Methods for regioselective N‐methylation and ‐alkylation of unsaturated heterocycles with "off the shelf" reagents are highly sought‐after. This reaction could drastically simplify synthesis of privileged bioactive molecules. Here we report engineered and natural methyltransferases for challenging N‐(m)ethylation of heterocycles, including benzimidazoles, benzotriazoles, imidazoles and indazoles. The reactions are performed through a cyclic enzyme cascade that consists of two methyltransferases using only iodoalkanes or methyl tosylate as simple reagents. This method enables the selective synthesis of important molecules that are otherwise difficult to access, proceeds with high regioselectivity (r.r. up to >99 %), yield (up to 99 %), on a preparative scale, and with nearly equimolar concentrations of simple starting materials. [ABSTRACT FROM AUTHOR]

Biocatalysis has traditionally been viewed as a field that primarily enables access to chiral centers. This includes the synthesis of chiral alcohols, amines and carbonyl compounds, often through functional group interconversion via hydrolytic or oxidation‐reduction reactions. This limitation is partly being overcome by the design and evolution of new enzymes. Here, we provide an overview of a recently thriving research field that we summarize as biocatalytic alkylation chemistry. In the past 3–4 years, numerous new enzymes have been developed that catalyze sp3 C−C/N/O/S bond formations. These enzymes utilize different mechanisms to generate molecular complexity by coupling simple fragments with high activity and selectivity. In many cases, the engineered enzymes perform reactions that are difficult or impossible to achieve with current small‐molecule catalysts such as organocatalysts and transition‐metal complexes. This review further highlights that the design of new enzyme function is particularly successful when off‐the‐shelf synthetic reagents are utilized to access non‐natural reactive intermediates. This underscores how biocatalysis is gradually moving to a field that build molecules through selective bond forming reactions. [ABSTRACT FROM AUTHOR]

Copyright of Angewandte Chemie is the property of Wiley-Blackwell and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Selective alkylation of pyrazoles could solve a challenge in chemistry and streamline synthesis of important molecules. Here we report catalyst‐controlled pyrazole alkylation by a cyclic two‐enzyme cascade. In this enzymatic system, a promiscuous enzyme uses haloalkanes as precursors to generate non‐natural analogs of the common cosubstrate S‐adenosyl‐l‐methionine. A second engineered enzyme transfers the alkyl group in highly selective C−N bond formations to the pyrazole substrate. The cosubstrate is recycled and only used in catalytic amounts. Key is a computational enzyme‐library design tool that converted a promiscuous methyltransferase into a small enzyme family of pyrazole‐alkylating enzymes in one round of mutagenesis and screening. With this enzymatic system, pyrazole alkylation (methylation, ethylation, propylation) was achieved with unprecedented regioselectivity (>99 %), regiodivergence, and in a first example on preparative scale. [ABSTRACT FROM AUTHOR]

The direct enantioselective addition of water to unactivated alkenes could simplify the synthesis of chiral alcohols and solve a long‐standing challenge in catalysis. Here we report that an engineered fatty acid hydratase can catalyze the asymmetric hydration of various terminal and internal alkenes. In the presence of a carboxylic acid decoy molecule for activation of the oleate hydratase from E. meningoseptica, asymmetric hydration of unactivated alkenes was achieved with up to 93 % conversion, excellent selectivity (>99 % ee, >95 % regioselectivity), and on a preparative scale. Engineered for success: Asymmetric hydration of unactivated, aliphatic alkenes was achieved with a fatty acid hydratase generating chiral alcohols with high conversion, excellent regio‐ and enantioselectivity, and on a preparative scale. Key for high activity is a carboxylic acid decoy molecule that activates the wildtype and engineered enzymes for alkene hydration on alternative substrates. [ABSTRACT FROM AUTHOR]

A variant with three changes in the amino acid sequence was able to produce methylated derivatives of linear terpenoids with high selectivity by C-methylation of an unactivated alkene. Cover Picture: Methylation of Unactivated Alkenes with Engineered Methyltransferases To Generate Non-natural Terpenoids (Angew. Keywords: Alkene Methylation; Biocatalysis; Late-Stage Modification; Methyltransferase; Terpenoids EN Alkene Methylation Biocatalysis Late-Stage Modification Methyltransferase Terpenoids 1 1 1 06/20/23 20230626 NES 230626 B A methyltransferase b from the green alga I Chlamydomonas reinhardtii i was identified and engineered for late-stage modifications of the carbon skeleton of terpenes. [Extracted from the article]

A variant with three changes in the amino acid sequence was able to produce methylated derivatives of linear terpenoids with high selectivity by C-methylation of an unactivated alkene. A variant with three changes in the amino acid sequence was able to produce methylated derivatives of linear terpenoids with high selectivity by C-methylation of an unactivated alkene. Keywords: Alken-Methylierung; Biokatalyse; Diversifizierung; Methyltransferase; Terpenoide DE Alken-Methylierung Biokatalyse Diversifizierung Methyltransferase Terpenoide 1 1 1 06/20/23 20230626 NES 230626 B A methyltransferase b from the green alga I Chlamydomonas reinhardtii i was identified and engineered for late-stage modifications of the carbon skeleton of terpenes. [Extracted from the article]

The squalene hopene cyclase from Alicyclobacillus acidocaldarius ( AacSHC) is a highly efficient enzyme catalyst for stereoselective Brønsted acid catalysis. We engineered AacSHC to catalyze the selective Prins cyclization of citronellal. Four active site variants were identified for the diastereoselective cyclization of ( S)-citronellal to stereoisomers (−)- iso-isopulegol, (+)-isopulegol and (−)- neo-isopulegol, respectively. The replacement of active site residues resulted in two triple variants that catalyzed the transformation of ( R)-citronellal to give the isomers (+)- neo-isopulegol and (−)-isopulegol with up to >99 % de, respectively. The newly designed library of functionally diverse active site geometries exhibits high selective control during citronellal cyclization, leading exclusively to a single diastereomer of the desired isopulegol. Whereas the cyclization of citronellal with chemical catalysts was observed to produce the isopulegol isomer with the lowest energy, the reaction with AacSHC variants proceeded with higher product selectivity. The results of this study show that variants of AacSHC are excellent catalysts for the highly selective formation of isopulegol stereoisomers. [ABSTRACT FROM AUTHOR]

Imine reductases (IREDs) have emerged as promising biocatalysts for the synthesis of chiral amines. In this study, the asymmetric imine reductase-catalyzed intermolecular reductive amination with NADPH as the hydrogen source was investigated. A highly chemo- and stereoselective imine reductase was applied for the reductive amination by using a panel of carbonyls with different amine nucleophiles. Primary and secondary amine products were generated in moderate to high yields with high enantiomeric excess values. The formation of the imine intermediate was studied between carbonyl substrates and methylamine in aqueous solution in the pH range of 4.0 to 9.0 by 1H NMR spectroscopy. We further measured the kinetics of the reductive amination of benzaldehyde with methylamine. This imine reductase-catalyzed approach constitutes a powerful and direct method for the synthesis of valuable amines under mild reaction conditions. [ABSTRACT FROM AUTHOR]

Polycyclizations constitute a cornerstone of chemistry and biology. Multicyclic scaffolds are generated by terpene cyclase enzymes in nature through a carbocationic polycyclization cascade of a prefolded polyisoprene backbone, for which electrostatic stabilization of transient carbocationic species is believed to drive catalysis. Computational studies and site-directed mutagenesis were used to assess the contribution of entropy to the polycyclization cascade catalyzed by the triterpene cyclase from A. acidocaldarius. Our results show that entropy contributes significantly to the rate enhancement through the release of water molecules through specific channels. A single rational point mutation that results in the disruption of one of these water channels decreased the entropic contribution to catalysis by 60 kcal mol−1. This work demonstrates that entropy is the key to enzyme-catalyzed polycyclizations, which are highly relevant in biology since 90 % of all natural products contain a cyclic subunit. [ABSTRACT FROM AUTHOR]

The use of enzymes as catalysts for the preparation of novel compounds has received steadily increasing attention over the past few years. High demands are placed on the identification of new biocatalysts for organic synthesis. The catalysis of more ambitious reactions reflects the high expectations of this field of research. Enzymes play an increasingly important role as biocatalysts in the synthesis of key intermediates for the pharmaceutical and chemical industry, and new enzymatic technologies and processes have been established. Enzymes are an important part of the spectrum of catalysts available for synthetic chemistry. The advantages and applications of the most recent and attractive biocatalysts-reductases, transaminases, ammonia lyases, epoxide hydrolases, and dehalogenases-will be discussed herein and exemplified by the syntheses of interesting compounds. [ABSTRACT FROM AUTHOR]

Die Verwendung von Enzymen als Katalysatoren zur Synthese neuartiger Verbindungen ist in den letzten Jahren zunehmend in den Fokus gerückt. Entsprechend hohe Erwartungen werden an die Entdeckung und Identifizierung neuer Biokatalysatoren für die organische Synthese gestellt. Dies spiegelt sich auch in der Komplexität der adressierten Reaktionen wider. Der Anwendungsbereich von Biokatalysatoren umfasst die Synthese wichtiger Zwischenprodukte für die pharmazeutische und chemische Industrie sowie die Entwicklung neuer enzymatischer Techniken und Prozesse. Enzyme sind ein wichtiger Teil des Spektrums an Katalysatoren, die der Synthesechemie zur Verfügung stehen. Die Vorteile und Anwendungsmöglichkeiten der neuesten und höchst vielversprechenden Generation von Biokatalysatoren - Reduktasen, Transaminasen, Ammoniak-Lyasen, Epoxidhydrolasen und Dehalogenasen - werden hier vorgestellt und anhand der Synthese von Schlüsselmolekülen beispielhaft diskutiert. [ABSTRACT FROM AUTHOR]

Review: recent advances in enzyme catalysis for the synthesis of challenging chemicals; 128 refs. [ABSTRACT FROM AUTHOR]