NAD(P)H-Dependent Enzymes for Reductive Amination: Active Site Description and Carbonyl-Containing Compound Spectrum Laurine Ducrot, Megan Bennett, Gideon Grogan, Carine Vergne-Vaxelaire To cite this version: Laurine Ducrot, Megan Bennett, Gideon Grogan, Carine Vergne-Vaxelaire. NAD(P)H-Dependent En- zymes for Reductive Amination: Active Site Description and Carbonyl-Containing Compound Spec- trum. Advanced Synthesis and Catalysis, Wiley-VCH Verlag, In press, 10.1002/adsc.202000870. hal-02945508 HAL Id: hal-02945508 https://hal.archives-ouvertes.fr/hal-02945508 Submitted on 22 Sep 2020 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. REVIEW DOI: 10.1002/adsc.201((will be filled in by the editorial staff)) NAD(P)H-Dependent Enzymes for Reductive Amination: Active Site Description and Carbonyl-Containing Compound Spectrum. Laurine Ducrot,a Megan Bennett,b Gideon Groganb and Carine Vergne- Vaxelairea* a Génomique Métabolique, Genoscope, Institut François Jacob, CEA, CNRS, Univ Evry, Université Paris- Saclay, 91057 Evry, France b York Structural Biology Laboratory, Department of Chemistry, University of York, Heslington, York, YO10 5DD, UK. Received: ((will be filled in by the editorial staff)) Abstract. The biocatalytic asymmetric synthesis of amines In this review we summarize the development of such from carbonyl compounds and amine precursors presents an enzymes from the engineering of amino acid dehydrogenases important advance in sustainable synthetic chemistry. (AADHs) and opine dehydrogenases (OpDHs) to become Oxidoreductases (ORs) that catalyze the NAD(P)H- amine dehydrogenases (AmDHs), which are active toward dependent reductive amination of carbonyl compounds ketones devoid of any requisite carboxylate and/or amine directly to amines using amine donors present advantages functions, through to the discovery of native AmDHs and complementary to those of amine transaminases (ATAs) with reductive aminases (RedAms), and the engineering of all of respect to selectivity, stability and substrate scope. Indeed these scaffolds for improved or altered activity. Structural some ORs accept alkyl and aryl amines as reaction partners and mechanistic studies have revealed similarities, but also enabling access to chiral secondary amine products that are differences in the determinants of substrate binding and not directly accessible using ATAs. Moreover, superior atom mechanism in the enzymes. The survey reveals that a economy can usually be achieved as no sacrificial amines are complementary approach to enzyme discovery that utilizes required as with ATAs. In recent years a number of ORs that both natural genetic resources and engineering can be apparently catalyze both imine formation and imine reduction combined to deliver biocatalysts that have significant in the reductive amination of carbonyls has been identified potential for the industrial synthesis of chiral amines. using structure informed protein engineering, sequence analysis from natural biodiversity and increasingly a mixture Keywords: Amines; reductive amination; amine of both. dehydrogenases; oxidoreductases; imine reductases been the focus of research by both academic and 1 Introduction industrial groups. One main advantage of biocatalysis is that enzymes are intrinsically chiral, and they are For environmental and societal reasons, there is therefore often able to differentiate between increasing pressure to utilize green processes and enantiomers of a racemic substrate and to impart high renewable materials in industry. In the context of stereoselectivity to a transformation. Given the chemical processes, enzyme-mediated importance of chiral amine moieties in Active transformations, i.e. biocatalysis, are now considered Pharmaceutical Ingredients (APIs) and as credible alternatives to conventional synthesis as, in agrochemicals,[4] classes of enzymes able to access this general, they operate under milder conditions and offer functionality with high process efficiencies suitable shorter synthetic routes to desired products.[1] Efforts for industry, have emerged in the last decades. A to study biocatalytic alternatives are mainly directed majority of amines made in industry are synthesized toward functions present in bulk chemicals produced using reductive amination, namely the reaction of a in high tonnage or high value-added products such as carbonyl-containing compound, i.e. ketone or pharmaceuticals.[2] Among these, aliphatic primary aldehyde, with ammonia or amine in the presence of a amines[3] and chiral secondary amines are some of the reductant, to form the corresponding amine with one most important compounds, and they have therefore or more substitutions. In the case of chiral amines, 1 either enantioselective synthesis, or additional steps of (AADHs) engineered to possess amine dehydrogenase resolution (enzyme-catalyzed by lipases or not) are activity (AmDHs), native amine dehydrogenases (nat- employed. Asymmetric reductive amination mainly AmDHs), imine reductases (IREDs) with activity consists of organometallic catalysis, with the use of toward carbonyl-containing substrates, reductive unsustainable transition metals and hydrogen gas, and aminases (RedAms), a subclass of IREDs, and organocatalytic approaches.[5] In a recent review, engineered opine dehydrogenases (OpDHs) (Figure Afanasyev et al. describe all the pharmaceutical drugs, 1). classified by therapeutic targets, that are formed using reductive amination reactions.[6] The equivalent enzymatic process was defined as one of the major challenges for biocatalysis in the pharmaceutical industry.[7] In recent years, a number of different enzymes that enable the asymmetric reductive amination of carbonyl substrates have emerged. In addition to ω-transaminases, which carry out the formal reductive amination of carbonyls using PLP/PMP cofactors and an amine donor,[8] NAD(P)H- dependent oxidoreductases[9] performing reductive aminations have become increasingly established as important biocatalysts for these reactions.[10] One major advantage of these enzymes is the access to various efficient methods available to recycle their nicotinamide cofactor,[11] including self-sufficient [12] hydride transfer processes. The availability of such Figure 1. Reactions catalyzed by the enzymes discussed in enzymes for chemists has increased significantly in this review. recent years and includes amino acid dehydrogenases Laurine Ducrot is a Ph. D student Megan Bennett is a PhD student at Paris-Saclay University at the University of York (France) in the Laboratory of working with Professor Gideon Biocatalysis, Bioremediation and Grogan on the characterization Synthetic Metabolism (UMR of amine dehydrogenases and 8030 Genomics Metabolics, their biocatalytic applications. Genoscope) working with Dr. She studied for her MBiochem Carine Vergne-Vaxelaire on the degree at York, which included discovery and improvement of a project working within the native amine dehydrogenases for York Structural Biology the synthesis of chiral amines Laboratory. Her interests using genomic and structural include structural biology and approaches. She graduated from the National Graduate the applications of enzyme engineering in biotechnology. School of Chemistry of Montpellier (France) after a 6- month long project at Amano Enzyme in the R&D Center of Gifu (Japan). Gideon Grogan is Professor of Carine Vergne-Vaxelaire Biochemistry at York University. obtained her Ph. D in 2006 at His research interests include the the University of Paris-Saclay discovery, application and (France) under the supervision structure-informed engineering of Dr. Ali Al-Mourabit working of enzymes with the potential for ((Author Portrait)) on the isolation of key nitrogen- ((Author industrial application, including containing natural products Portrait)) ligases, oxygenases and from marine sponges and their dehydrogenases for reduction of biomimetic synthesis. After one both C=O and C=N bonds. He is year as medicinal chemist at the author of over 130 Servier (France), she joined the publications in the area of French Alternative Energies applied biocatalysis and a book and Atomic Energy Commission (CEA) in the newly funded Laboratory of Biocatalysis, 2 ‘Practical Biotransformations’, a guide to enzyme Bioremediation and Synthetic Metabolism of the technology for organic chemists. Genoscope Unit UMR 8030 Genomics Metabolics. Her research interests are in the area of biocatalysis with particular emphasis on the discovery of new biocatalysts among biodiversity by genome-mining approaches mastered in the unit, and their applications as green tools for the synthesis of key building blocks. Her current main project is focused upon discovering native amine dehydrogenases for the production of amines and undertaking structural and biochemical studies thereof. In this review we focus on the application of NAD(P)H-dependent enzymes that have been used to either catalyze, or enable, the asymmetric reductive amination of a carbonyl group (excluding α-ketoacids) with an amine donor, and