Article

Characterization of a New Cold-adapted Lipase from Pseudomonas sp. TK-3

Graduate School of Science and Engineering, University of Toyama, 3190 Gofuku, Toyama, 930-8555, Japan, .
Applied biochemistry and biotechnology (Impact Factor: 1.74). 06/2012; 168(2):327-38. DOI: 10.1007/s12010-012-9776-7
Source: PubMed

ABSTRACT

A psychrotrophic Pseudomonas sp. TK-3 was isolated from dirty and cool stream water in Toyama, Japan from which we cloned and characterized the bacterial lipase LipTK-3. The sequenced DNA fragment contains an open reading frame of 1,428 bp that encoded a protein of 476 amino acids with an estimated molecular mass of 50,132 Da. The lipase showed high sequence similarity to those of subfamily Ι.3 lipase and had a conserved GXSXG motif around the catalytic Ser residue. Its optimal temperature was 20-25 °C, lower than in most other subfamily Ι.3 lipases. The lipase exhibited about 30 % of maximal activity at 5 °C. The optimal pH value was 8.0. The activity was strongly inhibited by EDTA and was highly dependent on Ca(2+). Tricaprylin and p-nitrophenyl caprylate were the most favorable substrates among the triglycerides and p-nitrophenyl esters, respectively. LipTK-3 also showed high activity towards natural substrates including edible vegetable oils and animal fats. Furthermore, LipTK-3 was very active and stable in the presence of several detergents, metal ions, and organic solvents. This cold-adapted lipase may prove useful for future applications.

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    • "Tricaprylin and p-nitrophenyl caprylate were the most favorable substrates of lipase from Pseudomonas sp. TK-3 [23]. An important property of protein catalysts is their activity in organic solvents used in organic synthesis reactions. "
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    ABSTRACT: A novel cold-active lipase gene encoding 294 amino acid residues was obtained from the Yersinia enterocolitica strain KM1. Sequence alignment and phylogenetic analysis revealed that this novel lipase is a new member of the bacterial lipase family I.1. The lipase shares the conserved GXSXG motif and catalytic triad Ser85-Asp239-His261. The recombinant protein LipA was solubly and heterogeneously expressed in Escherichia coli, purified by Ni-affinity chromatography, and then characterized. LipA was active over a broad range spanning 15-60°C with an optimum activity at 25°C and across a wide pH range from 5.0 to 11.0 with an optimum activity at pH 7.5. The molecular weight was estimated to be 34.2 KDa. The lipase could be activated by Mg(2+) and a low concentration (10%) of ethanol, dimethyl sulfoxide, methanol and acetonitrile, whereas it was strongly inhibited by Zn(2+), Cu(2+) and Mn(2+). This cold-active lipase may be a good candidate for detergents and biocatalysts at low temperature. Copyright © 2015. Published by Elsevier Inc.
    Full-text · Article · Aug 2015 · Protein Expression and Purification
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    • "CtroL4 22.6 14.7 16.6 18.0±1.6 CaLA 43.0 47.1 40.2 43.5±3.5 AflaL0 44.4 44.6 40.9 43.3±2.1 2011; Tanaka et al. 2012; Litantra et al. 2013 "
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    No preview · Article · Jan 2013 · Journal of the Chinese Cereals and Oils Association
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