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Identification and characterization of a thermostable and cobalt-dependent amidase from Burkholderia phytofirmans ZJB-15079 for efficient synthesis of ( R)-3,3,3-trifluoro-2-hydroxy-2-methylpropionic acid.
- Source :
-
Applied Microbiology & Biotechnology . Mar2017, Vol. 101 Issue 5, p1953-1964. 12p. - Publication Year :
- 2017
-
Abstract
- Enantiomerically pure 3,3,3-trifluoro-2-hydroxy-2-methylpropionic acids are important chiral building blocks for a series of pharmaceuticals. Here, a bacteria strain with 3,3,3-trifluoro-2-hydroxy-2-methylpropanamide-degrading ability was screened and identified as Burkholderia phytofirmans ZJB-15079, from which a novel amidase ( Bp-Ami) was cloned and demonstrated to be capable of kinetic resolution of rac-3,3,3-trifluoro-2-hydroxy-2-methylpropanamide to optically pure ( R)-3,3,3-trifluoro-2-hydroxy-2-methylpropionic acid. Phylogenetic analysis revealed that Bp-Ami was closely located to the acetamidase/formamidase (FmdA_AmdA) family, and it shared high homology with acetamidases. Bp-Ami was found to be the first cobalt-dependent FmdA_AmdA family amidase. The enzyme activity was significantly increased by 37.7-fold in the presence of 1 mM Co, with a specific activity of 753.5 U/mg, K value of 24.73 mM, and k /K value of 22.47 mM s. As an enzyme from mesophile, Bp-Ami exhibited extreme thermostability with a half-life of 47.93 h at 80 °C, which was even superior to other reported amidases from thermophiles. The whole cell catalysis of 200 g/L 3,3,3-trifluoro-2-hydroxy-2-methylpropanamide by Escherichia coli harboring Bp-Ami (5 g/L) resulted in 44 % yield and an enantiomeric excess ( ee ) of 95 % within 10 min ( E = 86). The high substrate tolerance, high specific activity, and extreme thermostability demonstrated the great potential of Bp-Ami for efficient biocatalytic synthesis of ( R)-3,3,3-trifluoro-2-hydroxy-2-methylpropionic acid. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 01757598
- Volume :
- 101
- Issue :
- 5
- Database :
- Academic Search Index
- Journal :
- Applied Microbiology & Biotechnology
- Publication Type :
- Academic Journal
- Accession number :
- 121264251
- Full Text :
- https://doi.org/10.1007/s00253-016-7921-x