Substitution of S-(beta-aminoethyl)-cysteine for active-site lysine of thermostable aspartate aminotransferase.
The active site lysyl residue (K239) of the thermostable aspartate aminotransferase [EC 2.6.1.1] was replaced by cysteinyl residue by means of site-directed mutagenesis. The K239C mutant enzyme obtained was catalytically inactive. The reaction of the cysteinyl residue of the K239C mutant enzyme with ethylenimine led to the formation of S-(beta-aminoethylcysteinyl (SAEC) residue. The K239SAEC mutant enzyme obtained showed about 25% of the activity of wild-type enzyme, and absorbed at 375 nm, which suggested the internal Schiff base formation.
Yoshimura
T
T
Laboratory of Microbial Biochemistry, Kyoto University.
Matsushima
Y
Y
Tanizawa
K
K
Sung
M H
MH
Yamauchi
T
T
Wakayama
M
M
Esaki
N
N
Soda
K
K
D000596:Amino Acids / Q000032:analysis; D001219:Aspartate Aminotransferases / Q000235:genetics*; D001388:Aziridines / Q000494:pharmacology; D001407:Bacillus / Q000201:enzymology; D001665:Binding Sites / Q000187:drug effects; D003545:Cysteine / Q000031:analogs & derivatives* / Q000235:genetics; D008239:Lysine / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D013053:Spectrophotometry
D000596:Amino Acids; D001388:Aziridines; C000204:S-2-aminoethyl cysteine; C033132:aziridine; D001219:Aspartate Aminotransferases; D008239:Lysine; D003545:Cysteine