Active rat aromatic-L-amino acid decarboxylase as a fusion protein in Escherichia coli.
The DNA sequence encoding rat aromatic-L-amino acid decarboxylase (AADC) was inserted into the Escherichia coli (E. coli) expression vector pMAL-c2. This clone produced a fusion protein able to catalyze the conversion of L-DOPA to dopamine. After purification and treatment of the fusion protein by factor Xa (FXa), an enzymatically active form of the enzyme resistant to FXa was isolated. It showed kinetic constants, Vmax, K(m) and enzymatic properties very similar to those obtained previously for the mammalian enzyme. This method for obtaining active AADC appears to be useful for initiating the study of the catalytic activity of this protein because it permitted the rapid isolation and the stabilization of an active form of the enzyme.
Jebai
F
F
Laboratoire de biochimie cellulaire, Collège de France, Paris.
Thibault
J
J
Krieger
M
M
D000818:Animals; D001142:Aromatic-L-Amino-Acid Decarboxylases / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology*; D015951:Factor Xa / Q000494:pharmacology; D005822:Genetic Vectors; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
D011993:Recombinant Fusion Proteins; D015951:Factor Xa; D001142:Aromatic-L-Amino-Acid Decarboxylases