title: Enhancement of the activity of l-aspartase from Escherichia coli W by directed evolution.
abstract: l-Aspartase was modified by directed evolution. After four rounds of error-prone PCR and three rounds of DNA shuffling, an evolved enzyme purified from the final round showed a 28-fold increased k(cat)/K(m) and 4.6-fold decreased K(m). The thermostability and stable pH range were also enhanced. The DNA sequence of the evolved aspartase gene showed seven base changes, resulting in three amino acid changes from the native enzyme: N217K, T233R, V367G. The mechanism of the enhancement of activity was analyzed.
authors.affiliation: Key Laboratory of Molecular Enzymology and Engineering, Jilin University, Changchun, 130023, People's Republic of China.
authors.forename: L J
authors.initials: LJ
authors.lastname: Wang
authors.forename: X D
authors.initials: XD
authors.lastname: Kong
authors.forename: H Y
authors.initials: HY
authors.lastname: Zhang
authors.forename: X P
authors.initials: XP
authors.lastname: Wang
authors.forename: J
authors.initials: J
authors.lastname: Zhang
mesh_terms: D001220:Aspartate Ammonia-Lyase / Q000737:chemistry* / Q000235:genetics*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics*; D004926:Escherichia coli; D019143:Evolution, Molecular; D016133:Polymerase Chain Reaction; D017510:Protein Folding
chemical_list: D001426:Bacterial Proteins; D001220:Aspartate Ammonia-Lyase