title: Activation of aspartase by site-directed mutagenesis.
abstract: To elucidate the role of sulfhydryl groups in the enzymatic reaction of the aspartase from Escherichia coli, we used site-directed mutagenesis which showed that the enzyme was activated by replacement of Cys-430 with a tryptophan. This mutation produced functional alterations without appreciable structural change: The kcat values became 3-fold at pH 6.0; the Hill coefficient values became higher under both pH conditions; the dependence of enzyme activity on divalent metal ions increased; and hydroxylamine, a good substrate for the wild-type enzyme, proved a poor substrate for the mutant.
authors.affiliation: Department of Chemistry, Faculty of Science, Kyoto University, Japan.
authors.forename: S
authors.initials: S
authors.lastname: Murase
authors.forename: J S
authors.initials: JS
authors.lastname: Takagi
authors.forename: Y
authors.initials: Y
authors.lastname: Higashi
authors.forename: H
authors.initials: H
authors.lastname: Imaishi
authors.forename: N
authors.initials: N
authors.lastname: Yumoto
authors.forename: M
authors.initials: M
authors.lastname: Tokushige
mesh_terms: D000595:Amino Acid Sequence; D001220:Aspartate Ammonia-Lyase / Q000235:genetics / Q000378:metabolism*; D002942:Circular Dichroism; D003545:Cysteine; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments / Q000302:isolation & purification; D011487:Protein Conformation; D014364:Tryptophan
chemical_list: D010446:Peptide Fragments; D014364:Tryptophan; D001220:Aspartate Ammonia-Lyase; D003545:Cysteine