Angewandte Reviews Chemie How to cite: Enzyme Catalysis International Edition: doi.org/10.1002/anie.202006648 German Edition: doi.org/10.1002/ange.202006648 Biocatalysis: Enzymatic Synthesis for Industrial Appli- cations Shuke Wu, Radka Snajdrova, Jeffrey C. Moore, Kai Baldenius,* and Uwe T. Bornscheuer* Keywords: biocatalysis · enzyme catalysis · industrial catalysis · organic synthesis · stereoselectivity Angewandte Chemie &&&& 2020 The Authors. Published by Wiley-VCH GmbH Angew. Chem. Int. Ed. 2020, 59,2–34 Ü Ü These are not the final page numbers! Angewandte Reviews Chemie Biocatalysis has found numerous applications in various fields as an From the Contents alternative to chemical catalysis. The use of enzymes in organic synthesis, especially to make chiral compounds for pharmaceuticals as 1. Introduction 3 well for the flavors and fragrance industry, are the most prominent 2. Alcohols 5 examples. In addition, biocatalysts are used on a large scale to make specialty and even bulk chemicals. This review intends to give illus- 3. Amines 9 trative examples in this field with a special focus on scalable chemical production using enzymes. It also discusses the opportunities and 4. Carbonyls, Carboxylic Acids and Derivatives 12 limitations of enzymatic syntheses using distinct examples and provides an outlook on emerging enzyme classes. 5. Glycosylation 18 6. Complex Molecules 21 1. Introduction 7. Biocatalytic Reactions for Biocatalysis has developed in the last two decades into Potential Industrialization 23 a rather mature and widely used technology.[1] With a few noticeable exceptions, biocatalysis in the early 2000s kept 8. Conclusion and Outlook 26 hiding in niche applications and focused on the synthesis or resolution of optically active intermediates.[2] Since then, biocatalysis has evolved more and more into a broadly applicable tool for chemical synthesis and manufacturing as Here, the NHase from Rhodococcus rhodochrous J1—used documented in many books.[3] Important driving forces are in a whole-cell system to avoid enzyme isolation as no the rapid discovery of new enzyme variants by modern undesired side reactions occur and stability is higher— bioinformatics and computer modelling supported enzyme exhibits outstanding catalytic efficiency as up to 7 kg acryl- engineering.[4] While the tremendous catalytic activity of amide can be produced per gram cells with product concen- enzymes is widely recognized, often their stability and cost are trations exceeding 500 g per liter reactor volume and space- considered a limitation. In this review we will focus on time-yields (STY) exceeding 0.1 kgLÀ1 hÀ1. biocatalysis suitable for scalable chemical production and In the meantime, many more enzymes made it into large discuss the opportunities and limitations of enzymatic syn- scale biocatalytic processes for which several examples are theses using distinct examples. given in this review. One reason is faster and straight forward A search in Scopus (Elsevier) for reviews on “Biocatal- discovery and engineering of suitable biocatalysts (the 3rd[1a] ysis” reveals more than 2000 hits; in SciFinder (Chemical and 4th[6] “wave”). This includes access to a plethora of novel Abstract Service, CAS) with a less stringent definition of enzymes via protein sequence and structure databases, their “review” more than 5000 articles are documented. Our aim shall not be to add yet another review simply summarizing the [*] Dr. S. Wu, Prof. Dr. U. T. Bornscheuer latest achievements in the biocatalysis field. Instead, we Institute of Biochemistry, Dept. of Biotechnology & Enzyme rather intend to give guidance to synthetic chemists which Catalysis, Greifswald University biocatalytic conversion technology may serve his/her manu- Felix-Hausdorff-Strasse 4, 17487 Greifswald (Germany) facturing challenge best. For this purpose, important key E-mail: [email protected] Homepage: http://biotech.uni-greifswald.de/ performance indicators (KPIs) will be applied to provide efficiency considerations that qualify new biocatalytic pro- Dr. R. Snajdrova Novartis Institutes for BioMedical Research, Global Discovery cesses for industrial scale-up and commercialization. While Chemistry, 4056 Basel (Switzerland) we will give reference to more specialized reviews of the Dr. J. C. Moore individual biotransformations, our comprehensive approach Process Research and Development, Merck & Co., Inc. shall help synthetic chemists navigate to the most efficient 126 E. Lincoln Ave, Rahway, NJ 07065 (USA) route for a multistep synthesis involving biocatalysis. Dr. K. Baldenius When in the early 2000s seminal reviews appeared,[2] Baldenius Biotech Consulting biocatalysis was still mostly using hydrolases (such as lipase Hafenstr. 31, 68159 Mannheim (Germany) CAL-B) or amidases (such as penicillin acylase and Subtili- E-mail: [email protected] sin), predominantly for the kinetic resolution of chiral Homepage: http://www.baldenius-biotech.com primary and secondary alcohols, amines or carboxylic acids. The ORCID identification numbers for the authors of this article can Ketoreductases (KREDs, as a subgroup of alcohol dehydro- be found under https://doi.org/10.1002/anie.202006648. genases, ADHs) were employed to make chiral secondary 2020 The Authors. Published by Wiley-VCH GmbH. This is an open access article under the terms of the Creative Commons alcohols via asymmetric reduction of prochiral ketones. Attribution Non-Commercial NoDerivs License, which permits use Probably the most prominent large scale industrial biocata- and distribution in any medium, provided the original work is lytic process was the long-established nitrile hydratase properly cited, the use is non-commercial, and no modifications or (NHase) process to make acrylamide from acrylonitrile.[5] adaptations are made. Angew. Chem. Int. Ed. 2020, 59, 2 – 34 2020 The Authors. Published by Wiley-VCH GmbH www.angewandte.org &&&& These are not the final page numbers! Ü Ü Angewandte Reviews Chemie improvement guided by bioinformatic tools in combination Still, not every new biocatalytic reaction (theoretically) with rational design or directed evolution, high-throughput possible or working on small scale makes it into an industrial screening tools as well as a range of design methods as process for various reasons, as also pointed out by Hauer very summarized in reviews.[7] Especially directed evolution rep- recently.[11] Many of these reasons also apply to new chemical resents a key technology for which Frances H. Arnold was reactions, which never make it into production. For instance, awarded the Nobel prize in Chemistry in 2018.[8] The major it can be difficult to get a new process implemented simply acceleration of biocatalyst development in recent years stems because this requires new investments into a factory while an from the cheap availability of synthetic genes that allow for old process in a depreciated production site is still running rapid, affordable screening of a diverse set of enzyme variants. profitably. Furthermore, despite the achievements made in In addition, the strategic planning of enzymatic routes has enzyme discovery and engineering, the “need for speed” can been facilitated as several reviews[9] and a book[10] now cover still be an issue, as timelines for biocatalyst development retrosynthesis concepts for biocatalysis, which should ease the especially in the pharmaceutical industry are often very short decision of which type of enzyme (class) and reaction is most as stated in an excellent recent publication.[12] Other aspects suitable for a targeted product. Also, the combination of are given in Table 1. On the other hand, biocatalytic reactions biocatalysis with chemical catalysis (metal-, organo-, photo-, have the advantage that no special equipment is required and electro-catalysis) became more mature in the past decade.[3b] Shuke Wu obtained his PhD from the Kai Baldenius studied chemistry in Hamburg National University of Singapore (2015) and Southampton. He received his PhD for with Prof. Zhi Li (NUS) and Prof. Daniel I. research in asymmetric organometallic catal- C. Wang (MIT). In 2017, he moved to ysis, supervised by H. tom Dieck and H. B. Switzerland to work with Prof. Thomas R. Kagan. After his postdoc on natural product Ward at the University of Basel sponsored by synthesis with K. C. Nicolaou at the Scripps a Swiss government excellence scholarship. Research Institute he joined BASF in 1993. Currently, he is working with Prof. Uwe T. Kai served BASF in various functions (R&D, Bornscheuer at the University of Greifswald production, marketing, sales) before he took as an Alexander von Humboldt fellow. the lead of BASF’s biocatalysis research for almost a defcade. He left BASF to become a free-lancing consultant in 2019 and in 2020 he has founded Baldenius Biotech Consulting. Radka Snajdrova obtained her PhD from Uwe T. Bornscheuer studied chemistry and Vienna University of Technology in 2007, received his PhD in 1993 at Hannover followed by postdoctoral studies at York University followed by a postdoc at Nagoya University (UK) and Greifswald University University (Japan). In 1998, he completed (Germany). In 2010, she transitioned to his Habilitation at Stuttgart University industry applying and developing biocatalytic about the use of lipases and esterases in technologies at Novacta in the UK, prior to organic synthesis. He has been Professor at joining Chemical Process Development at the Institute of Biochemistry at Greifswald GSK, with responsibility for