title: A novel cold-adapted lipase from Acinetobacter sp. XMZ-26: gene cloning and characterisation.
abstract: Acinetobacter sp. XMZ-26 (ACCC 05422) was isolated from soil samples obtained from glaciers in Xinjiang Province, China. The partial nucleotide sequence of a lipase gene was obtained by touchdown PCR using degenerate primers designed based on the conserved domains of cold-adapted lipases. Subsequently, a complete gene sequence encoding a 317 amino acid polypeptide was identified. Our novel lipase gene, lipA, was overexpressed in Escherichia coli. The recombinant protein (LipA) was purified by Ni-affinity chromatography, and then deeply characterised. The LipA resulted to hydrolyse pNP esters of fatty acids with acyl chain length from C2 to C16, and the preferred substrate was pNP octanoate showing a k(cat) = 560.52 ± 28.32 s(-1), K(m) = 0.075 ± 0.008 mM, and a k(cat)/K(m) = 7,377.29 ± 118.88 s(-1) mM(-1). Maximal LipA activity was observed at a temperature of 15°C and pH 10.0 using pNP decanoate as substrate. That LipA peaked at such a low temperature and remained most activity between 5°C and 35°C indicated that it was a cold-adapted enzyme. Remarkably, this lipase retained much of its activity in the presence of commercial detergents and organic solvents, including Ninol, Triton X-100, methanol, PEG-600, and DMSO. This cold-adapted lipase may find applications in the detergent industry and organic synthesis.
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, Beijing 100081, People's Republic of China. xiaomeizheng84@yahoo.cn
authors.forename: Xiaomei
authors.initials: X
authors.lastname: Zheng
authors.forename: Xiaoyu
authors.initials: X
authors.lastname: Chu
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.forename: Ningfeng
authors.initials: N
authors.lastname: Wu
authors.forename: Yunliu
authors.initials: Y
authors.lastname: Fan
mesh_terms: D000150:Acinetobacter / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D004795:Enzyme Stability; D007700:Kinetics; D008049:Lipase / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D012988:Soil Microbiology; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D008049:Lipase