Production of D-p-hydroxyphenylglycine by N-carbamoyl-D-amino acid amidohydrolase-overproducing Escherichia coli strains.
The N-carbamoyl-D-amino acid amidohydrolase (D-carbamoylase) gene from Agrobacterium radiobacter NRRL B11291 has been successfully cloned and expressed in Escherichia coli. Subcloning of the D-carbamoylase gene into different types of vectors and backgrounds of E. coli strains showed that the optimal expression level of D-carbamoylase was achieved in a ColE1-derived plasmid with a 150-fold increase in specific enzyme activity compared to that in a pSC101-derived plasmid. In addition, the recombinant plasmids were very stable in the E. coli strain ATCC11303 but not in JCL1258 tested here. Employing the recombinant E. coli strain DH5alpha/pAH61 for D-p-hydroxyphenylglycine production showed that the cell was capable of transforming N-carbamoyl-D-hydroxylphenylglycine to D-p-hydroxyphenylglycine with a molar conversion yield of 100% and a production rate of 1.9 g/(L h). In comparison with A. radiobacter NRRL B11291, this productivity approximates a 55-fold increase in D-hydroxyphenylglycine production. This result suggests the potential application of recombinant E. coli strains for the transformation reaction.
Chao
Y P
YP
Department of Chemical Engineering, Feng Chia University, 100 Wenhwa Road, Taichung, Taiwan, and Department of Chemical Engineering, National Taiwan University of Science and Technology, Taipei 106, Taiwan.
Juang
T Y
TY
Chern
J T
JT
Lee
C K
CK
D000581:Amidohydrolases / Q000096:biosynthesis* / Q000235:genetics; D019149:Bioreactors; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism*; D005998:Glycine / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011995:Recombination, Genetic; D012231:Rhizobium / Q000201:enzymology / Q000235:genetics
C118766:N-carbamoyl-4-hydroxyphenylglycine; C023010:4-hydroxyphenylglycine; D000581:Amidohydrolases; C080477:N-carbamoyl-D-amino acid amidohydrolase; D005998:Glycine