Asymmetric synthesis of (R)-3-fluoroalanine from 3-fluoropyruvate using omega-transaminase

Abstract

In this study, (R)-3-fluoroalanine was asymmetrically synthesized from 3-fluoropyruvate (F-pyruvate) and (S)-α-methylbenzylamine (MBA) using recombinant ω-transaminase (TA) from Vibrio fluvialis JS17. The reaction was severely inhibited by acetophenone (deaminated product of α-MBA). In the presence of 5 mM acetophenone, the reactivity of the enzyme towards F-pyruvate decreased by 78%. To overcome the product inhibition by acetophenone, a biphasic reaction was successfully used. The conversion of F-pyruvate into (R)-3-fluoroalanine (enatiomeric exess (e.e.) > 99%) was about 95% in the biphasic system (75 mM F-pyruvate, 100 mM (S)-α-MBA, and 3.0 U/mL), whereas 31% was obtained without product extraction. The use of racemic α-MBA as an amino donor instead of (S)-α-MBA can reduce the reaction cost and also produce chiral amines through kinetic resolution. When the kinetic resolution of racemic α-MBA (40 mM) was carried out with F-pyruvate (30 mM) and ω-TA (3.0 U/mL) in 100 mM phosphate buffer (pH 7.0), the e.e. of (R)-α-MBA reached 98.4% with 52.2% conversion for 10 h and 21 mM (R)-3-fluoroalanine was produced with 70% conversion and an e.e. > 99%.

Chemoenzymatic Platform for Synthesis of Chiral Organofluorines Based on Type II Aldolases

“…From the initial b-fluoro-aketoacid fluoropyruvate adducts,w ed emonstrate the feasibility of two enzymatic transformations inspired by the metabolism of pyruvate in nature:d ehydrogenation and transamination. In fact, commercial lactate dehydrogenase, which we used earlier in stereochemical experiments, accepted 3c-l as substrates to afford the ß-fluoro-a-hydrox- yacids 9c-l.W ef urther show that the transaminase from Vibrio fluvialis (VfAT) [14] can convert 3h and 3j into the corresponding b-fluoro amino acids 10 h and 10 j,respectively. Finally,t op roduce 2'-o r3 '-fluorosugars,w es ubjected the previously purified acids or esters syn-4a, anti-4a, anti-4b, 6, and 7 to synthetic routes involving chemical reduction.…”

New Generation of Biocatalysts for Organic Synthesis

“…(R)-Fluoroalanine was obtained with 70 % conversion and > 99 % enantiomeric excess by using this biphasic system. [84]…”

Transaminases and their Applications

“…3-Fluoropyruvate + methylbenzylamine [47] l-Homoalanine Pyruvate + (rac)-β-amino-N-butyric acid KR [52] l-Isoleucine l-Alanine TA 2-Keto-3-methyl-valerate + l-alanine AS [53] the correct conformation has to be produced. Recently, the asymmetric synthesis of prochiral diketones to 3-substituted cyclohexylamine derivatives serving as a build ing block for the multidrug-resistant protein MRP1 has been introduced [58].…”

“…An approach for the asymmetric synthesis of (R)-fluoroalanine using a biphasic reaction to overcome product inhibition caused by acetophenone was developed by Bea et al [47]. The substrate fluoropyruvate and the product (R)-fluoroalanine remained in the aqueous phase, whereas acetophenone was extracted by isooctane, the selected organic solvent.…”