The Industrial Age of Biocatalytic Transamination Fuchs, Michael; Farnberger, Judith E.; Kroutil, Wolfgang
European journal of organic chemistry,
November 2015, Letnik:
2015, Številka:
32
Journal Article
Recenzirano
Odprti dostop
During the last decade the use of ω‐transaminases has been identified as a very powerful method for the preparation of optically pure amines from the corresponding ketones. Their immense potential ...for the preparation of chiral amines, together with their ease of use in combination with existing biocatalytic methods, have made these biocatalysts a competitor to any chemical methodology for (asymmetric) amination. An increasing number of examples, especially from industry, shows that this biocatalytic technology outmaneuvers existing chemical processes by its simple and flexible nature. In the last few years numerous publications and patents on synthetic routes, mainly to pharmaceuticals, involving ω‐transaminases have been published. The review gives an overview of the application of ω‐transaminases in organic synthesis with a focus on active pharmaceutical ingredients (APIs) and the developments during the last few years.
ω‐Transaminase‐catalyzed preparation of chiral amines has emerged as one of the most successful fields in biocatalysis over the past ten years. This review covers the development of the catalytic systems and points out the most important applications, especially with regard to industrial and commercial use of the technology.
The cleavage of aryl methyl ethers is a common reaction in chemistry requiring rather harsh conditions; consequently, it is prone to undesired reactions and lacks regioselectivity. Nevertheless, ...O-demethylation of aryl methyl ethers is a tool to valorize natural and pharmaceutical compounds by deprotecting reactive hydroxyl moieties. Various oxidative enzymes are known to catalyze this reaction at the expense of molecular oxygen, which may lead in the case of phenols/catechols to undesired side reactions (e.g., oxidation, polymerization). Here an oxygen-independent demethylation via methyl transfer is presented employing a cobalamin-dependent veratrol-O-demethylase (vdmB). The biocatalytic demethylation transforms a variety of aryl methyl ethers with two functional methoxy moieties either in 1,2-position or in 1,3-position. Biocatalytic reactions enabled, for instance, the regioselective monodemethylation of substituted 3,4-dimethoxy phenol as well as the monodemethylation of 1,3,5-trimethoxybenzene. The methyltransferase vdmB was also successfully applied for the regioselective demethylation of natural compounds such as papaverine and rac-yatein. The approach presented here represents an alternative to chemical and enzymatic demethylation concepts and allows performing regioselective demethylation in the absence of oxygen under mild conditions, representing a valuable extension of the synthetic repertoire to modify pharmaceuticals and diversify natural products.
Isomerization is a fundamental reaction in chemistry. However, isomerization of phenyl methyl ethers has not been described yet. Using a cobalamin-dependent methyl transferase, a reversible shuttle ...concept was investigated for isomerization of catechol monomethyl ethers. The methyl ether of substituted catechol derivatives was successfully transferred onto the adjacent hydroxy moiety. For instance, the cobalamin-dependent biocatalyst transformed isovanillin to its regioisomer vanillin with significant regioisomeric excess (68% vanillin). To the best of our knowledge, isomerization by methyl transfer employing a methyl transferase has not been reported before.
Regioselective reactions represent a significant challenge for organic chemistry. Here the regioselective methylation of a single hydroxy group of 4‐substituted catechols was investigated employing ...the cobalamin‐dependent methyltransferase from Desulfitobacterium hafniense. Catechols substituted in position four were methylated either in meta‐ or para‐position to the substituent depending whether the substituent was polar or apolar. While the biocatalytic cobalamin dependent methylation was meta‐selective with 4‐substituted catechols bearing hydrophilic groups, it was para‐selective for hydrophobic substituents. Furthermore, the presence of water miscible co‐solvents had a clear improving influence, whereby THF turned out to enable the formation of a single regioisomer in selected cases. Finally, it was found that also the pH led to an enhancement of regioselectivity for the cases investigated.
Biocatalysis: Regioselective reactions represent a challenge for organic chemistry. Here the regioselective mono‐methylation of 4‐substituted catechols was investigated employing the cobalamin‐dependent methyltransferase from Desulfitobacterium hafniense. The influence of polar and apolar substituents in position four, co‐solvents, as well as pH were examined. Water miscible co‐solvents allowed to tune the regioselectivity, whereby THF turned out to enable the formation of a single regioisomer in selected cases.
The ether functionality represents a very common motif in organic chemistry and especially the methyl ether is commonly found in natural products. Its formation and cleavage can be achieved via ...countless chemical procedures. Nevertheless, since in particular the cleavage often involves harsh reaction conditions, milder alternatives are highly demanded. Very recently, we have reported on a biocatalytic shuttle catalysis concept for reversible cleavage and formation of phenolic O‐methyl ethers employing a corrinoid‐dependent methyl transferase system from the anaerobic organism Desulfitobacterium hafniense. Here we report the technical study of this system, focusing on the demethylation of guaiacol as model reaction. The optimal buffer‐, pH‐, temperature‐ and cofactor‐preferences were determined as well as the influence of organic co‐solvents. Beside methyl cobalamin also hydroxocobalamin turned out to be a suitable cofactor species, although the latter required activation. Various O‐methyl phenyl ethers were successfully demethylated with conversions up to 82% at 10 mM substrate concentration.
Transaminases have become a key tool in biocatalysis to introduce the amine functionality into a range of molecules like prochiral α-ketoacids and ketones. However, due to the necessity of shifting ...the equilibrium towards the product side (depending on the amine donor) an efficient amination system may require three enzymes. So far, this well-established transformation has mainly been performed in vitro by assembling all biocatalysts individually, which comes along with elaborate and costly preparation steps. We present the design and characterization of a flexible approach enabling a quick set-up of single-cell biocatalysts producing the desired enzymes. By choosing an appropriate co-expression strategy, a modular system was obtained, allowing for flexible plug-and-play combination of enzymes chosen from the toolbox of available transaminases and/or recycling enzymes tailored for the desired application.
By using a two-plasmid strategy for the recycling enzyme and the transaminase together with chromosomal integration of an amino acid dehydrogenase, two enzyme modules could individually be selected and combined with specifically tailored E. coli strains. Various plug-and-play combinations of the enzymes led to the construction of a series of single-cell catalysts suitable for the amination of various types of substrates. On the one hand the fermentative amination of α-ketoacids coupled both with metabolic and non-metabolic cofactor regeneration was studied, giving access to the corresponding α-amino acids in up to 96% conversion. On the other hand, biocatalysts were employed in a non-metabolic, "in vitro-type" asymmetric reductive amination of the prochiral ketone 4-phenyl-2-butanone, yielding the amine in good conversion (77%) and excellent stereoselectivity (ee = 98%).
The described modularized concept enables the construction of tailored single-cell catalysts which provide all required enzymes for asymmetric reductive amination in a flexible fashion, representing a more efficient approach for the production of chiral amines and amino acids.
Abstract
Synthetically established methods for methylation of phenols and demethylation of methyl phenyl ethers rely in general on hazardous reagents or/and harsh reaction conditions and are ...irreversible. Consequently, alternative regioselective methods for the reversible formation and breakage of C-O-ether bonds to be performed under mild and sustainable conditions are highly desired. Here we present a biocatalytic shuttle concept making use of corrinoid-dependent methyl transferases from anaerobic bacteria. The two-component enzymatic system consists of a corrinoid protein carrying the cofactor and acting as methyl group shuttle, and a methyltransferase catalyzing both methylation and demethylation in a reversible fashion. Various phenyl methyl ethers are successfully demethylated and serve in addition as sustainable methylating agents for the functionalization of various substituted catechols. Therefore, this methyl transfer approach represents a promising alternative to common chemical protocols and a valuable add-on for the toolbox of available biocatalysts.
Over the last decades biocatalysis has emerged as an indispensable and versatile tool for the asymmetric synthesis of active pharmaceutical ingredients (APIs). In this context, especially ...transaminases (TAs) have been successfully used for the preparation of numerous α‐chiral, optically pure amines, serving as important building blocks for APIs. Here we elaborate on the development of transaminases recognizing the α‐chiral centre adjacent to an aldehyde moiety with aliphatic residues, opening up concepts for novel synthetic routes to the antiepileptic drugs Brivaracetam and Pregabalin. The transformation proceeded via dynamic kinetic resolution (DKR) based on the bio‐induced racemisation of the aldehyde enantiomers, enabling the amination of the racemic substrates with quantitative conversions. Medium, substrate as well as enzyme engineering gave access to both (R)‐ and (S)‐enantiomers of the amine precursors of the stereocomplementary drugs in high optical purity, representing a short route to mentioned APIs.
Over the last decades biocatalysis has emerged as an indispensable and versatile tool for the asymmetric synthesis of active pharmaceutical ingredients (APIs). In this context, especially ...transaminases (TAs) have been successfully used for the preparation of numerous alpha-chiral, optically pure amines, serving as important building blocks for APIs. Here we elaborate on the development of transaminases recognizing the alpha-chiral centre adjacent to an aldehyde moiety with aliphatic residues, opening up concepts for novel synthetic routes to the antiepileptic drugs Brivaracetam and Pregabalin. The transformation proceeded via dynamic kinetic resolution (DKR) based on the bio-induced racemisation of the aldehyde enantiomers, enabling the amination of the racemic substrates with quantitative conversions. Medium, substrate as well as enzyme engineering gave access to both (R)- and (S)-enantiomers of the amine precursors of the stereocomplementary drugs in high optical purity, representing a short route to mentioned APIs.
Various enantiocomplementary ω-transaminases (ωTAs) were investigated in kinetic resolution and asymmetric reductive amination reactions to prepare silodosin amine. Whilst the enzymatic kinetic ...resolution gave moderate to good results with respect to the yield and enantioselectivity, the asymmetric reductive amination proved to be superior. The best results were obtained with the ωTA originating from (R)-Arthrobacter sp. which afforded the desired bioactive (R)-enantiomer in enantiomerically pure form (ee >97%) at excellent conversion (conv. >97%) under mild and benign reaction conditions.
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