In this review the current state-of-the-art of S-adenosylmethionine (SAM)-dependent methyltransferases and SAM are evaluated. Their structural classification and diversity is introduced and key mechanistic aspects presented which are then detailed further. Then, catalytic SAM as a target for drugs, and approaches to utilise SAM as a cofactor in synthesis are introduced with different supply and regeneration approaches evaluated. The use of SAM analogues are also described. Finally O-, N-, C- and S-MTs, their synthetic applications and potential for compound diversification is given.@en

Biocatalysis is one of the most promising technologies for the sustainable synthesis of molecules for pharmaceutical, biotechnological and industrial purposes. From the gram to the ton scale, biocatalysis is employed with success. This is underpinned by the fact that the global enzyme market is predicted to increase from $7 billion to $10 billion by 2024. This review concentrates on showing the strong benefits that biocatalysis and the use of enzymes can provide to synthetic chemistry. Several examples of successful implementations of enzymes are discussed highlighting not only high-value pharmaceutical processes but also low-cost bulk products. Thus, biocatalytic methods make the chemistry more environmentally friendly and product specific.@en

S-Adenosyl-l-methionine (SAM)-dependent methyltransferases (MTs) are highly chemoselective enzymes grouped in C-, N-, O-, S- and halide MTs, depending on the (hetero) atom that acts as the methyl group acceptor. So far, OMTs present the largest group, including many well investigated candidates. The catechol OMT from mammals such as from Rattus norvegicus (RnCOMT) is involved in the metabolism of neurotransmitters like dopamine. It is known to methylate the hydroxyl of the catechol ring in the 3 position. There are also reports showing that the regioselectivity of different COMTs can vary leading to different products with methyl groups in the 3 and or 4 positions. Nevertheless, there was only O-methylation reported for COMTs. Another related MT, the caffeate OMT involved in the lignin biosynthesis of plants has also been reported as a chemoselective enzyme. In nature, S-methylation is a rare phenomenon with different methyl donors being involved in the methyl transfer onto sulfur atoms. Several SAM-dependent MTs are identified as S-methyltransferases (SMTs), these are involved in salvaging pathways and xenobiotic metabolism of cells. Here, we report a new function of three OMTs; RnCOMT, a COMT from Myxococcus xanthus (MxSafC), and a CaOMT from Prunus persica (PpCaOMT) with acceptance towards different aromatic thiol substrates with up to full conversion.@en

In nature 2-deoxy-D-ribose-5-phosphate aldolase (DERA) catalyses the reversible formation of 2-deoxyribose 5-phosphate from D-glyceraldehyde 3-phosphate and acetaldehyde. In addition, this enzyme can use acetaldehyde as the sole substrate, resulting in a tandem aldol reaction, yielding 2,4,6-trideoxy-D-erythro-hexapyranose, which spontaneously cyclizes. This reaction is very useful for the synthesis of the side chain of statin-type drugs used to decrease cholesterol levels in blood. One of the main challenges in the use of DERA in industrial processes, where high substrate loads are needed to achieve the desired productivity, is its inactivation by high acetaldehyde concentration. In this work, the utility of different variants of Pectobacterium atrosepticum DERA (PaDERA) as whole cell biocatalysts to synthesize 2-deoxyribose 5-phosphate and 2,4,6-trideoxy-D-erythro-hexapyranose was analysed. Under optimized conditions, E. coli BL21 (PaDERA C-His AA C49M) whole cells yields 99 % of both products. Furthermore, this enzyme is able to tolerate 500 mM acetaldehyde in a whole-cell experiment which makes it suitable for industrial applications.@en

© 2018, The Author(s). 2-Deoxy-d-ribose-5-phosphate aldolase (DERA) is a class I aldolase that offers access to several building blocks for organic synthesis. It catalyzes the stereoselective C–C bond formation between acetaldehyde and numerous other aldehydes. However, the practical application of DERA as a biocatalyst is limited by its poor tolerance towards industrially relevant concentrations of aldehydes, in particular acetaldehyde. Therefore, the development of proper experimental conditions, including protein engineering and/or immobilization on appropriate supports, is required. The present review is aimed to provide a brief overview of DERA, its history, and progress made in understanding the functioning of the enzyme. Furthermore, the current understanding regarding aldehyde resistance of DERA and the various optimizations carried out to modify this property are discussed.@en

The implementation of a stereoselective Michael addition with water as substrate is still a major challenge by classical, chemical means. Inspired by nature's ability to carry out this attractive reaction with both high selectivity and efficiency, the interest in hydratases (EC 4.2.1.x) to accomplish a selective water addition is steadily rising. The gram-positive bacterial genus Rhodococcus is known as biocatalytic powerhouse and has been reported to hydrate various Michael acceptors leading to chiral alcohols. This study aimed at the in-depth re-investigation of the hydration potential of Rhodococcus whole-cells towards Michael acceptors. Here, two concurrent effects responsible for the hydration reaction were found: while the majority of substrates was hydrated in an oxygen-independent manner by amino-acid catalysis, an enzyme-catalysed water addition to (E)-4-hydroxy-3-methylbut-2-enoic acid was proven to be oxygen-dependent. 18O2-labelling studies showed that no 18O2 was incorporated in the product. Therefore, a novel O2-dependent hydratase distinct from all characterised hydratases so far was found.@en

DERA (2-Deoxy-D-ribose 5-phosphate aldolase) is the only known aldolase that accepts two aldehyde substrates, which makes it an attractive catalyst for the synthesis of a chiral polyol motif that is present in several pharmaceuticals, such as atorvastatin and pravastatin. However, inactivation of the enzyme in the presence of aldehydes hinders its practical application. Whole cells of Pectobacterium atrosepticum were reported to exhibit good tolerance toward acetaldehyde and to afford 2-deoxyribose 5-phosphate with good yields. The DERA gene (PaDERA) was identified, and both the wild-type and a C49M mutant were heterologously expressed in Escherichia coli. The purification protocol was optimized and an initial biochemical characterization was conducted. Unlike other DERAs, which show a maximal activity between pH 4.0 and 7.5, PaDERA presented an optimum pH in the alkaline range between 8.0 and 9.0. This could warrant its use for specific syntheses in the future. PaDERA also displayed fourfold higher specific activity than DERA from E. coli (EcDERA) and displayed a promising acetaldehyde resistance outside the whole-cell environment. The C49M mutation, which was previously identified to increase acetaldehyde tolerance in EcDERA, also led to significant improvements in the acetaldehyde tolerance of PaDERA.@en

This study presents a three-step one pot enzymatic cascade for the synthesis of a δ-lactone. Utilising acetaldehyde, combining 2-deoxyribose-5-phosphate aldolase (DERA) with an alcohol dehydrogenase (ADH) and a cofactor regeneration system this δ-lactone is synthesised with the same stereochemistry as the statin side chain precursor. The initial stage in this cascade involves the double aldol reaction, catalysed by DERA to produce the chiral lactone precursor from the achiral substrate acetaldehyde. The main challenge at this stage is the instability of DERA in the presence of high acetaldehyde concentrations. Therefore, Lactobacillus brevis DERA with a high natural acetaldehyde tolerance was genetically engineered to further improve this property. LbDERA C42M E78K exhibited improved activity and stability (no activity loss over 2 h) compared to the wild type (20% activity loss). In the second stage of the cascade, the aldol product is selectively oxidised to the lactone. A commercially available ADH was identified to selectively catalyse this oxidation using NADP+ as electron acceptor. NADP+ regeneration was achieved using O2 as substrate in two different ways: using either photo-activated flavin or NADPH oxidase (NOX). The lactone was successfully purified from the enzymatic cascades from a preparative scale reaction in 97% purity with an optical rotation [α]D = +34.2° (c = 0.7), proving the feasibility in a multi-enzyme three-step one-pot cascade.@en