title: Cloning, expression in Escherichia coli, and enzymatic properties of laccase from Aeromonas hydrophila WL-11.
abstract: A strain WL-11 with high laccase activity was isolated from activated sludge collected from the effluent treatment plant of a textile and dyeing industry. It was identified as Aeromonas hydrophila by physiological test and 16S rDNA sequence analysis. A gene encoding of laccase from a newly isolated Aeromonas hydrophila WL-11 was cloned and characterized. Nucleotide sequence analysis showed an open reading frame of 1605 bp encoding a polypeptide comprised of 534 amino acids. The primary structure of the enzyme predicted the structural features characteristic of other laccases, including the conserved regions of four histidine-rich copper-binding sites. The predicted amino acid sequence showed a high homology (more than 60%) with bacterial laccases in the genome and protein databases and the highest degree of similarity (61% identity) was observed with the multicopper oxidase of Klebsiella sp. 601. When expressed in Escherichia coli, the recombinant enzyme was overproduced in the cytoplasm as soluble and active form. The purified enzyme had an optimum pH of 2.6 and 8.0 for ABTS (2,2'-azino-bis(3-ethylbenzthiazolinesulfonic acid) and DMP (2,6-dimethoxyphenol), respectively. The kinetic study on ABTS revealed a higher affinity of this enzyme to this substrate than DMP.
authors.affiliation: Department of Biotechnology, Dong-A University, Busan 604-714, South Korea.
authors.forename: Jing
authors.initials: J
authors.lastname: Wu
authors.forename: Kyoung-Sook
authors.initials: KS
authors.lastname: Kim
authors.forename: Jai-Heon
authors.initials: JH
authors.lastname: Lee
authors.forename: Young-Choon
authors.initials: YC
authors.lastname: Lee
mesh_terms: D016980:Aeromonas hydrophila / Q000145:classification / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D042845:Laccase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012722:Sewage / Q000382:microbiology
chemical_list: D012722:Sewage; D042845:Laccase