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Monoacylated diamines are common motifs present in pharmaceuticals, agrochemicals, and natural products. However, the synthesis of these compounds often requires selective protection/deprotection s...

Sustainable production of fine chemicals and biofuels from renewable biomass offers a potential alternative to the continued use of finite geological oil reserves. However, in order to compete with current petrochemical refinery processes, alternative biorefinery processes must overcome significant costs and productivity barriers. Herein, we demonstrate the biocatalytic production of the versatile chemical building block, coniferol, for the first time, directly from lignocellulosic biomass. Following the biocatalytic treatment of lignocellulose to release and convert ferulic acid with feruloyl esterase (XynZ), carboxylic acid reductase (CAR) and aldo-keto reductase (AKR), this whole cell catalytic cascade not only achieved equivalent release of ferulic acid from lignocellulose compared to alkaline hydrolysis, but also displayed efficient conversion of ferulic acid to coniferol. This system represents a consolidated biodegradation–biotransformation strategy for the production of high value fine chemicals from waste plant biomass, offering the potential to minimize environmental waste and add value to agro-industrial residues.\ud \ud

The alkylation of amines with either alcohols or carboxylic acids represents a mild and safe alternative to the use of genotoxic alkyl halides and sulfonate esters. Here we report two complementary one-pot systems in which the reductive aminase (RedAm) from Aspergillus oryzae is combined with either (i) a 1° alcohol/alcohol oxidase (AO) or (ii) carboxylic acid/carboxylic acid reductase (CAR) to affect N-alkylation reactions. The application of both approaches has been exemplified with respect to substrate scope and also preparative scale synthesis. These new biocatalytic methods address issues facing alternative traditional synthetic protocols such as harsh conditions, overalkylation and complicated workup procedures.

Carboxylic acid reductases (CARs) are an emerging biocatalyst platform for the synthesis of a diverse array of aldehydes from carboxylic acids, operating chemoselectively and under mild aqueous conditions. As such, there is growing interest in the industrial application of these enzymes, both for the synthesis of aldehyde end-products, which are particularly prevalent in the flavor and fragrance industries, and aldehyde intermediates in multi-enzyme cascades. This perspective aims to review recent developments in the applications of CARs with a focus on the challenges and considerations involved in their implementation, as well as potential solutions with a view to increased industrial utility.

Fatty amine synthesis from renewable sources is an energetically-demanding process involving toxic metal catalysts and harsh reaction conditions as well as selectivity problems. Herein we present a mild, biocatalytic alternative to the conventional amination of fatty acids through a one-pot tandem cascade performed by a carboxylic acid reductase (CAR) and a transaminase (ω-TA). Saturated and unsaturated fatty acids, with carbon chain lengths ranging from C6 to C18, were successfully aminated obtaining conversions of up to 96%.

Carboxylic acid reductases (CARs) catalyze the reduction of a broad range of carboxylic acids to aldehydes using the cofactors adenosine triphosphate and nicotinamide adenine dinucleotide phosphate, and have become attractive biocatalysts for organic synthesis. Mechanistic understanding of CARs was used to expand reaction scope, generating biocatalysts for amide bond formation from carboxylic acid and amine. CARs demonstrated amidation activity for various acids and amines. Optimization of reaction conditions, with respect to pH and temperature, allowed for the synthesis of the anticonvulsant ilepcimide with up to 96 % conversion. Mechanistic studies using site-directed mutagenesis suggest that, following initial enzymatic adenylation of substrates, amidation of the carboxylic acid proceeds by direct reaction of the acyl adenylate with amine nucleophiles.

Carboxylic acid reductase (CAR) catalyzes the ATP- and NADPH-dependent reduction of carboxylic acids to the corresponding aldehydes. The enzyme is related to the nonribosomal peptide synthetases, consisting of an adenylation domain fused via a peptidyl carrier protein (PCP) to a reductase termination domain. Crystal structures of the CAR adenylation-PCP didomain demonstrate that large-scale domain motions occur between the adenylation and thiolation states. Crystal structures of the PCP-reductase didomain reveal that phosphopantetheine binding alters the orientation of a key Asp, resulting in a productive orientation of the bound nicotinamide. This ensures that further reduction of the aldehyde product does not occur. Combining crystallography with small-angle X-ray scattering (SAXS), we propose that molecular interactions between initiation and termination domains are limited to competing PCP docking sites. This theory is supported by the fact that (R)-pantetheine can support CAR activity for mixtures of the isolated domains. Our model suggests directions for further development of CAR as a biocatalyst.

Correction for ‘A biocatalytic cascade for the conversion of fatty acids to fatty amines’ by Joan Citoler, et al., Green Chem., 2019, 21, 4932–4935.

The application of biocatalysis in the pharmaceutical industry is rapidly growing as a result of increased access to enzymes that meet the demands of industrial processes. This expansion of activity has led to a corresponding increase in the demand for immobilised enzymes. EziG™ (EnginZyme AB, Sweden) is marketed as a general matrix for enzyme immobilisation on controlled porosity glass. In this work we identified criteria for a “general” enzyme immobilisation technology in the context of the requirements of the pharmaceutical industry. We subsequently evaluated EziG™ for generality in a series of case studies for the application of immobilised biocatalysts. In this study we have focussed on the challenges facing both academic and industrial applications such as enzyme stability, multistep reactions and reactions in continuous flow.

The direct C–H carboxylation of aromatic compounds is an attractive route to the corresponding carboxylic acids, but remains challenging under mild conditions. It has been proposed that the first step in anaerobic microbial degradation of recalcitrant aromatic compounds is a UbiD-mediated carboxylation. In this study, we use the UbiD enzyme ferulic acid decarboxylase (Fdc) in combination with a carboxylic acid reductase to create aromatic degradation-inspired cascade reactions, leading to efficient functionalization of styrene through CO2 fixation. We reveal that rational structure-guided laboratory evolution can expand the substrate scope of Fdc, resulting in activity on a range of mono- and bicyclic aromatic compounds through a single mutation. Selected variants demonstrated 150-fold improvement in the conversion of coumarillic acid to benzofuran + CO2 and unlocked reactivity towards naphthoic acid. Our data demonstrate that UbiD-mediated C–H activation is a versatile tool for the transformation of aryl/alkene compounds and CO2 into commodity chemicals. Biocatalytic cascade reactions using engineered variants of ferulic acid decarboxylase coupled to carboxylic acid reductase utilize carbon dioxide fixation to enable the carboxylation and functionalization of styrene and other aromatic compounds.