Engineered Enzyme Cascades for the Synthesis of Chiral Amines, Amides, and Nitriles

Francesco G. Mutti
Van ’t Hoff Institute for Molecular Sciences, HIMS-Biocat, University of Amsterdam

Abstract
The biocatalytic synthesis of chiral amines offers numerous advantages over chemocatalytic methods in terms of efficiency, selectivity, environmental sustainability, and applicability to diverse substrates. [1] In this context, my group has focused on the synthesis of chiral amines from prochiral ketones using ω-transaminases (ωTAs), as well as alternative novel aminating enzymes such as amine dehydrogenases (AmDHs) and imine reductases (IReds).[2]
We have created a new family of AmDHs based on the enzyme scaffold of an ε-deaminating L-lysine dehydrogenase (LysEDH). The best variant, LE-AmDH-v1, was highly thermostable, retained its catalytic activity after incubation at up to 50 °C for several days, and was applied to the synthesis of pharmaceutically relevant amines in enantiopure form.[3]
AmDHs, IReds, and ωTAs were also incorporated into biocatalytic cascades with other enzyme families for the conversion of alcohols, styrene derivatives, α-amino acids, or α,β-unsaturated ketones into chiral amines or amino alcohols containing up to two stereogenic centers.[4]
One example is the biocatalytic conversion of alcohols into α-chiral amines (i.e., a hydride-borrowing cascade) using alcohol dehydrogenases (ADHs) and an AmDH; we developed this biotransformation using isolated enzymes,[5] co-immobilized enzymes,[6] or E. coli cells in vivo.[7] The concept was extended to the synthesis of amino alcohols with two stereogenic centers.[8] 
During these studies on biocatalytic alcohol oxidation, we also observed an unprecedented conversion of alcohols into nitriles catalyzed by an alcohol oxidase variant, using only ammonia and dioxygen from air as reagents.[9] More recently, we extended this cascade to the synthesis of amides.

Selected references
[1] E. Abdelraheem, et al. Biocatalytic Amine Synthesis, in Comprehensive Chirality (Ed.: J. Cossy), Elsevier, 2024, pp. 210-304.
[2] a) T. Knaus, et al., Green Chem. 2017, 19, 453-463; b) W. Böhmer, et al., Adv. Synth. Catal. 2020, 362, 1858-1867.
[3] V. Tseliou, et al., Nat. Commun. 2019, 10, 3717.
[4] a) T. Knaus, et al., ACS Catal 2022, 12, 14459-14475; b) M. L. Corrado, et al., Org. Process Res. Dev. 2022, 26, 2085-2095.
[5] a) F. G. Mutti, et al., Science 2015, 349, 1525-1529; b) V. Tseliou, et al., Chem. Eur. J. 2021, 27, 3315-3325.
[6] W. Böhmer, et al., ChemCatChem 2018, 10, 731-735.
[7] J. A. Houwman, et al., Green Chem. 2019, 21, 3846-3857.
[8] M. L. Corrado, et al., Green Chem. 2019, 21, 6246-6251.
[9] J. Vilim, et al., Angew. Chem. Int. Ed. 2018, 57, 14240-14244.

Francesco Mutti is a Professor of Biocatalysis at the University of Amsterdam (Netherlands). He graduated in Industrial Chemistry (2004) and obtained his PhD in Chemistry (2008) from the University of Milan (Italy), under the supervision of Prof. Michele Gullotti, while also spending one year of his PhD in the group of Prof. Wolfgang Kroutil at the University of Graz.
After obtaining his PhD, he moved back to Graz as a Research Associate in the group of Prof. Wolfgang Kroutil at the University of Graz (2009–2012).
He then joined the group of Prof. Nicholas Turner at the University of Manchester (UK; 2013–2014).
Since 2015, he has been the Group Leader of Biocatalysis at the University of Amsterdam.
Among other achievements, he has been a Marie Skłodowska-Curie fellow and has received an ERC Starting Grant, as well as several EU grants, Dutch national grants, and industry funding. He has published 75 research papers, 4 book chapters, 9 book sections, and 7 world patents, and he has delivered 57 lectures at international conferences and companies.
His main research interests include the development of biocatalytic cascades for the sustainable synthesis of chemicals, enzyme discovery and engineering, continuous-flow biocatalysis, in vivo biocatalysis, bioelectrochemistry and bio-photocatalysis.

> Flyer