title: Construction of a chimeric aminopeptidase by a combination of gene shuffling and mutagenesis.
abstract: Three chimeric genes were constructed by gene shuffling of aminopeptidases from Aeromonas caviae and Vibrio proteolyticus. Although expressed chimeric enzymes formed inclusion bodies in Escherichia coli, the introduction of two amino acid mutations into the chimeric genes by site-saturated mutagenesis and a random mutation on error-prone PCR resulted in solubilization of the chimeric enzyme. In addition, active chimeric enzyme showed a different thermostability and thermoactivity to parental enzymes.
authors.affiliation: Enzyme Laboratory, National Food Research Institute, National Agriculture and Food Research Organization, Tsukuba, Ibaraki 305-8642, Japan. stnirasa@affrc.go.jp
authors.forename: Satoru
authors.initials: S
authors.lastname: Nirasawa
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Hayashi
mesh_terms: D000595:Amino Acid Sequence; D000626:Aminopeptidases / Q000737:chemistry / Q000235:genetics*; D032621:DNA Shuffling / Q000379:methods*; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics*; D016415:Sequence Alignment; D012995:Solubility
chemical_list: D011994:Recombinant Proteins; D000626:Aminopeptidases