Computer Modeling Explains the Structural Reasons for the Difference in Reactivity of Amine Transaminases Regarding Prochiral Methylketones

Amine transaminases (ATAs) are pyridoxal-5′-phosphate (PLP)-dependent enzymes that catalyze the transfer of an amino group from an amino donor to an aldehyde and/or ketone. In the past decade, the enzymatic reductive amination of prochiral ketones catalyzed by ATAs has attracted the attention of researchers, and more traditional chemical routes were replaced by enzymatic ones in industrial manufacturing. In the present work, the influence of the presence of an α,β-unsaturated system in a methylketone model substrate was investigated, using a set of five wild-type ATAs, the (R)-selective from Aspergillus terreus (Atr-TA) and Mycobacterium vanbaalenii (Mva-TA), the (S)-selective from Chromobacterium violaceum (Cvi-TA), Ruegeria pomeroyi (Rpo-TA), V. fluvialis (Vfl-TA) and an engineered variant of V. fluvialis (ATA-256 from Codexis). The high conversion rate (80 to 99%) and optical purity (78 to 99% ee) of both (R)- and (S)-ATAs for the substrate 1-phenyl-3-butanone, using isopropylamine (IPA) as an amino donor, were observed. However, the double bond in the α,β-position of 4-phenylbut-3-en-2-one dramatically reduced wild-type ATA reactivity, leading to conversions of <10% (without affecting the enantioselectivity). In contrast, the commercially engineered V. fluvialis variant, ATA-256, still enabled an 87% conversion, yielding a corresponding amine with >99% ee. Computational docking simulations showed the differences in orientation and intermolecular interactions in the active sites, providing insights to rationalize the observed experimental results.

Recent Advance in Organocatalyzed Asymmetric Reduction of Prochiral Ketones: An Update

Chiral alcohols are important synthetic intermediates or building blocks for the diverse synthesis of drugs, agrochemicals, and natural products. Asymmetric reduction of prochiral ketones has been the most popularly investigated method for accessing chiral alcohols. In this regard, the organocatalyzed asymmetric reduction as a complementary of transition-metal- and enzyme-catalyzed reactions have attracted tremendous interest in the past decades due to the nature of metal-free and easy operation, as well as, principly, the ease of recovery and reuse of catalysts. Following up a comprehensive overview on organocatalyzed asymmetric reduction of prochiral ketones in early 2018, this short review is intended to summarize the recent progress in this area from the beginning of the year 2018 to the end of Aug. 2021.

Chiral Synthesis of 3-Amino-1-phenylbutane by a Multi-Enzymatic Cascade System

Asymmetric synthesis of chiral amines from prochiral ketones using transaminases is an attractive biocatalytic strategy. Nevertheless, it is hampered by its unfavorable thermodynamic equilibrium. In the present work, an insitu by-product removal strategy was applied for the synthesis of 3-amino-1-phenylbutane (3-APB) by coupling a transaminase with a pyruvate decarboxylase (PDC), which does not require the use of any expensive additional cofactor. Using this strategy, the pyruvate obtained in the transamination reaction is transformed by PDC into acetaldehyde and CO2 which are of high volatility. Two different transaminases from Chromobacterium violaceum (CviTA) and Vibrio fluvialis (VflTA) were characterized to find out the appropriate pH conditions. In both cases, the addition of PDC dramatically enhanced 3-APB synthesis. Afterwards, different reaction conditions were tested to improve reaction conversion and yield. It was concluded that 30 °C and a 20-fold alanine excess lead to the best process metrics. Under the mentioned conditions, yields higher than 60% were reached with nearly 90% selectivity using both CviTA and VflTA. Moreover, high stereoselectivity for (S)-3-APB was obtained and ee of around 90% was achieved in both cases. For the first time, the asymmetric synthesis of 3-APB using PDC as by-product removal system using CviTA is reported.

Altering the Stereoselectivity of Whole-Cell Biotransformations via the Physicochemical Parameters Impacting the Processes

The enantioselective synthesis of organic compounds is one of the great challenges in organic synthetic chemistry due to its importance for the acquisition of biologically active derivatives, e.g., pharmaceuticals, agrochemicals, and others. This is why biological systems are increasingly applied as tools for chiral compounds synthesis or modification. The use of whole cells of “wild-type” microorganisms is one possible approach, especially as some methods allow improving the conversion degrees and controlling the stereoselectivity of the reaction without the need to introduce changes at the genetic level. Simple manipulation of the culture conditions, the form of a biocatalyst, or the appropriate composition of the biotransformation medium makes it possible to obtain optically pure products in a cheap, safe, and environmentally friendly manner. This review contains selected examples of the influence of physicochemical factors on the stereochemistry of the biocatalytic preparation of enantiomerically pure compounds, which is undertaken through kinetically controlled separation of their racemic mixtures or reduction of prochiral ketones and has an effect on the final enantiomeric purity and enantioselectivity of the reaction.

N-Phenylputrescine (NPP): A Natural Product Inspired Amine Donor for Biocatalysis

The synthesis of chiral amines in enantioenriched form is a keystone reaction in applied chemical synthesis. There is a strong push to develop greener and more sustainable alternatives to the metal-catalysed methods currently used in the pharmaceutical, agrochemical and fine chemical industries. A biocatalytic approach using transaminase (TA or ATA) enzymes to convert prochiral ketones to chiral amines with unparalleled levels of enantioselectivity is highly appealing. However, the use of TA enzymes in synthesis is severely hampered by the unfavourable thermodynamics associated with the amine donor/acceptor equilibrium. Several ‘smart’ amine donors have been developed that leverage chemical and physical driving forces to overcome this challenging equilibrium. Alongside this strategy, enzyme engineering is typically required to develop TAs compatible with these non-physiological amine donors and the unnatural reaction conditions they require. We herein disclose N-phenylputrescine (NPP) as a readily accessible amine donor, inspired by the biosynthesis of the dipyrroloquinoline alkaloids. NPP is compatible with a broad range of synthetically useful TA biocatalysts and performs across an unparalleled variety of reaction conditions (pH and temperature). Synthetic applicability has been demonstrated through the synthesis of the anti-diabetic drug sitagliptin, delivering the product in excellent enantiopurity using just two equivalents of NPP<br>

N-Phenylputrescine (NPP): A Natural Product Inspired Amine Donor for Biocatalysis

The synthesis of chiral amines in enantioenriched form is a keystone reaction in applied chemical synthesis. There is a strong push to develop greener and more sustainable alternatives to the metal-catalysed methods currently used in the pharmaceutical, agrochemical and fine chemical industries. A biocatalytic approach using transaminase (TA or ATA) enzymes to convert prochiral ketones to chiral amines with unparalleled levels of enantioselectivity is highly appealing. However, the use of TA enzymes in synthesis is severely hampered by the unfavourable thermodynamics associated with the amine donor/acceptor equilibrium. Several ‘smart’ amine donors have been developed that leverage chemical and physical driving forces to overcome this challenging equilibrium. Alongside this strategy, enzyme engineering is typically required to develop TAs compatible with these non-physiological amine donors and the unnatural reaction conditions they require. We herein disclose N-phenylputrescine (NPP) as a readily accessible amine donor, inspired by the biosynthesis of the dipyrroloquinoline alkaloids. NPP is compatible with a broad range of synthetically useful TA biocatalysts and performs across an unparalleled variety of reaction conditions (pH and temperature). Synthetic applicability has been demonstrated through the synthesis of the anti-diabetic drug sitagliptin, delivering the product in excellent enantiopurity using just two equivalents of NPP<br>

Characterisation of a Hyperthermophilic Transketolase from Thermotoga maritima DSM3109 as a Biocatalyst for 7-keto-Octuronic Acid Synthesis

Transketolase (TK) is a fundamentally important enzyme in industrial biocatalysis which carries out a stereospecific carbon-carbon bond formation, and is widely used in the synthesis of prochiral ketones. This study...

Continuous-flow stereoselective reduction of prochiral ketones in a whole cell bioreactor with natural deep eutectic solvents

Immobilized whole cells of Rhodotorula rubra MIM147 were used in a packed bed flow reactor for the enantioselective reduction of β-ketonitriles and for the efficient production of a key building...