Selection, Purification, Characterisation, and Cloning of a Novel Heat-Stable Stereo-Specific Amidase from Klebsiella o xytoca, and Its Application in the Synthesis of Enantiomerically Pure (R)- and (S)-3,3,3-Trifluoro-2-hydroxy-2-methylpropionic Acids and (S)-3,3,3-Trifluoro-2-hydroxy-2-methylpropionamide

• Published in: Organic process research & development Vol. 6; no. 4; pp. 497 - 504
• Main Authors: Shaw, Nicholas M, Naughton, Andrew, Robins, Karen, Tinschert, Andreas, Schmid, Evelyne, Hischier, Marie-Louise, Venetz, Veronika, Werlen, Josef, Zimmermann, Thomas, Brieden, Walter, de Riedmatten, Patricia, Roduit, Jean-Paul, Zimmermann, Bertin, Neumüller, Roman
• Format: Journal Article
• Language: English
• Published: American Chemical Society 19.07.2002
• ISSN: 1083-6160; 1520-586X
• DOI: 10.1021/op020025d

We isolated, characterised, and cloned an enantio-specific amidase from Klebsiella oxytoca and used it to resolve (R,S)-3,3,3-trifluoro-2-hydroxy-2-methylpropionamide, giving (R)-3,3,3-trifluoro-2-hydroxy-2-methylpropionic acid and (S)-3,3,3-trifluoro-2-hydroxy-2-methylpropionamide. The (S)-amide could then be hydrolysed chemically to (S)-3,3,3-trifluoro-2-hydroxy-2-methylpropionic acid. The process can therefore be adapted to produce both (R)- and (S)-enantiomers of 3,3,3-trifluoro-2-hydroxy-2-methylpropionic acid, or (S)-3,3,3-trifluoro-2-hydroxy-2-methylpropionamide. The biocatalytic step is part of a combined chemical and biocatalytic route that starts from ethyl trifluoroacetoacetate. The products typically have a purity of greater than 98% and ee values of essentially 100% after isolation. The process has been used to produce 100-g amounts of the (S)-acid, and successfully scaled up to produce 100-kg amounts of the (R)-acid, with the biotransformation carried out at the 1500-L scale.