Novel Old Yellow Enzyme Subclasses

Peters, Christin; Frasson, David; Sievers, Martin; Buller, Rebecca

doi:10.1002/cbic.201800770

Cited by 34 publications

(65 citation statements)

References 70 publications

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“…The enzyme Ppo-Er1 from P. polymyxa was discovered during the screening of 19 bacterial wild-type strains from the Culture Collection of Switzerland, as previously described [15]. Ppo-Er1 (41.3 kDa) is characterized by a substantial sequence similarity with the old yellow enzyme YqiG from Bacillus subtilis (50%) [34], Bac-OYE2 from Bacillus sp.…”

Section: Resultsmentioning

confidence: 99%

Section: Isolated In 1932 Bymentioning

confidence: 99%

“…Notably, amino acid sequence alignments of OYE homologs show high conservation in specific regions of the proteins, such as residues involved in catalysis, FMN, and substrate binding [7,15,23]. To account for these differences in sequence and the resulting structural features, the old yellow enzyme family can be further divided into five subclasses [15]. While enzyme members of the subclass I, also termed "classical" old yellow enzymes, and class II, introduced by Scrutton's group in 2010 and dubbed "thermophilic-like" [23], have been well explored [7,14], the recently described class III-V are less well investigated [15,24].…”

Section: Isolated In 1932 Bymentioning

confidence: 99%

“…Synthetic applications of ene reductases are manifold and range from the preparation of profens [25][26][27] and chiralγ-amino acids [28][29][30] to the synthesis of chiral phosphonates [31] and nitroalkanes [32], precursors in the synthesis of pharmaceutically active ingredients. To further promote an off-the-shelve synthetic use of ene reductases, which can reduce the time and cost of the implementation of a biocatalytic step into a process significantly, we set out to expand the available biocatalytic toolbox [15]. In this context, not only the discovery and engineering of novel ene reductases is of great utility [33], but also a careful characterization of the new biocatalysts is needed as it may lead to the construction of a more targeted enzyme library associated with reduced screening time and costs.…”

Section: Isolated In 1932 Bymentioning

confidence: 99%

“…While enoate reductases, medium-and short-chain dehydrogenases/reductases (MDR and SDR), as well as the recently discovered quinone Figure 1. Overview of the classification within the ene reductase family [15]. QnoR (NADPHdependent quinone reductase like ene-reductases), EnoR (enoate reductase), OYE (old yellow enzyme), MDR (medium-chain dehydrogenase/reductase), and SDR (short-chain dehydrogenase/reductase); Class I (classical OYE); Class II (thermophilic-like OYE) and Class V (fungal OYE).…”

Section: Introductionmentioning

confidence: 99%

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Characterization of the Novel Ene Reductase Ppo-Er1 from Paenibacillus Polymyxa

2020

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Ene reductases enable the asymmetric hydrogenation of activated alkenes allowing the manufacture of valuable chiral products. The enzymes complement existing metal- and organocatalytic approaches for the stereoselective reduction of activated C=C double bonds, and efforts to expand the biocatalytic toolbox with additional ene reductases are of high academic and industrial interest. Here, we present the characterization of a novel ene reductase from Paenibacillus polymyxa, named Ppo-Er1, belonging to the recently identified subgroup III of the old yellow enzyme family. The determination of substrate scope, solvent stability, temperature, and pH range of Ppo-Er1 is one of the first examples of a detailed biophysical characterization of a subgroup III enzyme. Notably, Ppo-Er1 possesses a wide temperature optimum (Topt: 20–45 °C) and retains high conversion rates of at least 70% even at 10 °C reaction temperature making it an interesting biocatalyst for the conversion of temperature-labile substrates. When assaying a set of different organic solvents to determine Ppo-Er1′s solvent tolerance, the ene reductase exhibited good performance in up to 40% cyclohexane as well as 20 vol% DMSO and ethanol. In summary, Ppo-Er1 exhibited activity for thirteen out of the nineteen investigated compounds, for ten of which Michaelis–Menten kinetics could be determined. The enzyme exhibited the highest specificity constant for maleimide with a kcat/KM value of 287 mM−1 s−1. In addition, Ppo-Er1 proved to be highly enantioselective for selected substrates with measured enantiomeric excess values of 92% or higher for 2-methyl-2-cyclohexenone, citral, and carvone.

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Section: Resultsmentioning

confidence: 99%

Section: Isolated In 1932 Bymentioning

confidence: 99%

Section: Isolated In 1932 Bymentioning

confidence: 99%

Section: Isolated In 1932 Bymentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Characterization of the Novel Ene Reductase Ppo-Er1 from Paenibacillus Polymyxa

2020

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Flavoprotein‐dependent Bioreduction

Toogood

Scrutton

2021

Flavin‐Based Catalysis

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Asymmetric Reduction of (R)‐Carvone through a Thermostable and Organic‐Solvent‐Tolerant Ene‐Reductase

et al. 2020

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Ene‐reductases allow regio‐ and stereoselective reduction of activated C=C double bonds at the expense of nicotinamide adenine dinucleotide cofactors [NAD(P)H]. Biological NAD(P)H can be replaced by synthetic mimics to facilitate enzyme screening and process optimization. The ene‐reductase FOYE‐1, originating from an acidophilic iron oxidizer, has been described as a promising candidate and is now being explored for applied biocatalysis. Biological and synthetic nicotinamide cofactors were evaluated to fuel FOYE‐1 to produce valuable compounds. A maximum activity of (319.7±3.2) U mg−1 with NADPH or of (206.7±3.4) U mg−1 with 1‐benzyl‐1,4‐dihydronicotinamide (BNAH) for the reduction of N‐methylmaleimide was observed at 30 °C. Notably, BNAH was found to be a promising reductant but exhibits poor solubility in water. Different organic solvents were therefore assayed: FOYE‐1 showed excellent performance in most systems with up to 20 vol% solvent and at temperatures up to 40 °C. Purification and application strategies were evaluated on a small scale to optimize the process. Finally, a 200 mL biotransformation of 750 mg (R)‐carvone afforded 495 mg of (2R,5R)‐dihydrocarvone (>95 % ee), demonstrating the simplicity of handling and application of FOYE‐1.

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Novel Old Yellow Enzyme Subclasses

Cited by 34 publications

References 70 publications

Characterization of the Novel Ene Reductase Ppo-Er1 from Paenibacillus Polymyxa

Characterization of the Novel Ene Reductase Ppo-Er1 from Paenibacillus Polymyxa

Flavoprotein‐dependent Bioreduction

Asymmetric Reduction of (R)‐Carvone through a Thermostable and Organic‐Solvent‐Tolerant Ene‐Reductase

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