Biocatalytic N-Alkylation of Amines Using Either Primary Alcohols or Carboxylic Acids via Reductive Aminase Cascades.

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.

Is it time for biocatalysis in fragment-based drug discovery?

The use of biocatalysts for fragment-based drug discovery has yet to be fully investigated, despite the promise enzymes hold for the synthesis of poly-functional, non-protected small molecules. Here we analyze products of the biocatalysis literature to demonstrate the potential for not only fragment generation, but also the enzyme-mediated elaboration of these fragments. Our analysis demonstrates that biocatalytic products can readily populate 3D chemical space, offering diverse catalytic approaches to help generate new, bioactive molecules.

Natural heterogeneous catalysis with immobilised oxidase biocatalysts.

The generation of immobilised oxidase biocatalysts allowing multifunctional oxidation of valuable chemicals using molecular oxygen is described. Engineered galactose oxidase (GOase) variants M1 and M3-5, an engineered choline oxidase (AcCO6) and monoamine oxidase (MAO-N D9) displayed long-term stability and reusability over several weeks when covalently attached on a solid support, outperforming their free counterparts in terms of stability (more than 20 fold), resistance to heat at 60 °C, and tolerance to neat organic solvents such as hexane and toluene. These robust heterogenous oxidation catalysts can be recovered after each reaction and be reused multiple times for the oxidation of different substrates.

Asymmetric synthesis of primary amines catalyzed by thermotolerant fungal reductive aminases.

Chiral primary amines are important intermediates in the synthesis of pharmaceutical compounds. Fungal reductive aminases (RedAms) are NADPH-dependent dehydrogenases that catalyse reductive amination of a range of ketones with short-chain primary amines supplied in an equimolar ratio to give corresponding secondary amines. Herein we describe structural and biochemical characterisation as well as synthetic applications of two RedAms from Neosartorya spp. (NfRedAm and NfisRedAm) that display a distinctive activity amongst fungal RedAms, namely a superior ability to use ammonia as the amine partner. Using these enzymes, we demonstrate the synthesis of a broad range of primary amines, with conversions up to >97% and excellent enantiomeric excess. Temperature dependent studies showed that these homologues also possess greater thermal stability compared to other enzymes within this family. Their synthetic applicability is further demonstrated by the production of several primary and secondary amines with turnover numbers (TN) up to 14 000 as well as continous flow reactions, obtaining chiral amines such as (R)-2-aminohexane in space time yields up to 8.1 g L-1 h-1. The remarkable features of NfRedAm and NfisRedAm highlight their potential for wider synthetic application as well as expanding the biocatalytic toolbox available for chiral amine synthesis.

Imine Reductases, Reductive Aminases, and Amine Oxidases for the Synthesis of Chiral Amines: Discovery, Characterization, and Synthetic Applications.

Synthesis of the chiral amine moiety is a key challenge for synthetic organic chemistry due to its prevalence in many biologically active molecules. Imine reductase and amine oxidase enzymes have enabled the biocatalytic synthesis of a host of chiral amine compounds. In this chapter, procedures for the synthesis of chiral amines using imine reductases (IREDs), the recently discovered IRED homologues reductive aminases, and amine oxidases (AOs) are described. Amine oxidases have been the subject of mutagenesis approaches for improvement of substrate scope. The high-throughput screening method for determining active variants in amine oxidase libraries is illustrated. Finally, in an approach which takes inspiration from nature, many enzymes can be combined with each other in cascade reactions. The incorporation of imine reductase and monoamine oxidase biocatalysts into several cascade reactions, both in vitro and in vivo (where the approach moves toward synthetic biology), is reported.

Imine reductases (IREDs).

Imine reductases (IREDs) have emerged as a valuable new set of biocatalysts for the asymmetric synthesis of optically active amines. The development of bioinformatics tools and searchable databases has led to the identification of a diverse range of new IRED biocatalysts that have been characterised and employed in different synthetic processes. This review describes the latest developments in the structural and mechanistic aspects of IREDs, together with synthetic applications of these enzymes, and identifies ongoing and future challenges in the field.