Molecular chaperones facilitate the soluble expression of N-acyl-D-amino acid amidohydrolases in Escherichia coli.
The overproduction of D-aminoacylase ( D-ANase, 233.8 U/mg), N-acyl-D-glutamate amidohydrolase (D-AGase, 38.1 U/mg) or N-acyl-D-aspartate amidohydrolase (D-AAase, 6.2 U/mg) in Escherichia coli is accompanied by aggregation of the overproduced protein. To facilitate the expression of active enzymes, the molecular chaperones GroEL-GroES (GroELS), DnaK-DnaJ-GrpE (DnaKJE), trigger factor (TF), GroELS and DnaKJE or GroELS and TF were coexpressed with the enzymes. D-ANase (313.3 U/mg) and D-AGase (95.8 U/mg) were overproduced in an active form at levels 1.3- and 1.8-fold higher, respectively, upon co-expression of GroELS and TF. An E. coli strain expressing the D-AAase gene simultaneously with the TF gene exhibited a 4.3-fold enhancement in d-AAase activity (32.0 U/mg) compared with control E. coli expressing the D-AAase gene alone.
Yoshimune
Kazuaki
K
Department of Applied Chemistry, Faculty of Engineering, Oita University, Dannoharu, Oita, 870-1192, Japan.
Ninomiya
Yoko
Y
Wakayama
Mamoru
M
Moriguchi
Mitsuaki
M
D000581:Amidohydrolases / Q000096:biosynthesis* / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000378:metabolism / Q000502:physiology*; D015964:Gene Expression Regulation, Bacterial; D005818:Genetic Engineering; D018832:Molecular Chaperones / Q000096:biosynthesis / Q000502:physiology*; D010957:Plasmids / Q000235:genetics
D029968:Escherichia coli Proteins; D018832:Molecular Chaperones; D000581:Amidohydrolases; C070988:N-acyl-D-glutamate deacylase