Clean Enzymatic Oxidation of 12α-Hydroxysteroids to 12-Oxo-Derivatives Catalyzed by Hydroxysteroid Dehydrogenase

Clean Enzymatic Oxidation of 12α-Hydroxysteroids to 12-Oxo-Derivatives Catalyzed by Hydroxysteroid Dehydrogenase

Tonin, F. (TU Delft BT/Biocatalysis)
Alvarenga Da Silva, N. (TU Delft BT/Biocatalysis)
Ye, Jia Zheng (Student TU Delft)
Arends, I.W.C.E. (TU Delft BT/Biocatalysis; Universiteit Utrecht)
Hanefeld, U. (TU Delft BT/Biocatalysis)

2019

The C12 specific oxidation of hydroxysteroids is an essential reaction required for the preparation of pharmaceutical ingredients like ursodeoxycholic acid (UDCA) and chenodeoxycholic acid (CDCA), which can be synthesized by Wolff-Kishner reduction of the obtained 12-oxo-hydroxysteroids. 12α-hydroxysteroid dehydrogenases (12α-HSDHs) have been shown to perform this reaction with high yields, under mild conditions and without the need of protection and deprotection steps, required in chemical synthesis. Here, the recombinant expression and biochemical characterization of the nicotinamide adenine dinucleotide (NAD ⁺ )-dependent HSDH from Eggerthella lenta (El12α-HSDH) are reported. This enzyme shows comparable properties with the well-known nicotinamide adenine dinucleotide phosphate (NADP ⁺ )-dependent enzyme from Clostridium sp. 48–50. In order to perform a viable and atom efficient enzymatic hydroxysteroid oxidation, NAD(P)H oxidase (NOX) was employed as cofactor regeneration system: NOX uses oxygen (O ₂ ) as sacrificial substrate and produces only water as side product. 10 mM of cholic acid was fully and selectively converted to 12-oxo-CDCA in 24 h. The possibility to employ this system on UCA and 7-oxo-deoxycholic acid (7-oxo-DCA) as substrates was additionally investigated. The performance of the El12α-HSDH was evaluated also in combination with a “classical” regeneration system (oxaloacetate/malate dehydrogenase) showing full conversion in 4 h. Finally, the feasibility of a catalytic aerobic-NAD ⁺ -dependent enzymatic oxidation was shown on a preparative scale (oxidation of CA to 12-oxo-CDCA) employing the El12α-HSDH-NOX system in a segmented-flow-reactor. (Figure presented.).

12α-hydroxysteroid dehydrogenases
Bile acids
Flow-reactor
NAD(P)H oxidase
NAD -dependent

http://resolver.tudelft.nl/uuid:3b0a4e8c-2533-42d1-bd30-0ca63c74e8bf

https://doi.org/10.1002/adsc.201900144

1615-4150

Advanced Synthesis & Catalysis, 361 (11), 2448-2455

Institutional Repository

journal article

© 2019 F. Tonin, N. Alvarenga Da Silva, Jia Zheng Ye, I.W.C.E. Arends, U. Hanefeld