Characterizing LipR from Pseudomonas sp. R0-14 and Applying in Enrichment of Polyunsaturated Fatty Acids from Algal Oil.

Journal of microbiology and biotechnology

PubMedID: 26215266

Yang W, Xu L, Zhang H, Yan Y. Characterizing LipR from Pseudomonas sp. R0-14 and Applying in Enrichment of Polyunsaturated Fatty Acids from Algal Oil. J Microbiol Biotechnol. 2015;.
In this study, Pseudomonas R0-14, which was isolated from Arctic soil samples, showed a clear halo when grown on M9 medium agarose plates containing olive oil-rhodamine B as substrate, suggesting that it expressed putative lipase(s). A putative lipase gene, lipR, was cloned from R0-14 by genome walking and Touchdown PCR. lipR encodes a 562-amino-acid polypeptide showing a typical a/ß hydrolase structure with a catalytic triad consisting of Ser153-Asp202-His260 and one a-helical lid (residues 103-113). A phylogenetic analysis revealed that LipR belongs to the lipase subfamily I. 3. LipR was successfully expressed in Escherichia coli, purified, and biochemically characterized. Recombinant LipR exhibited its maximum activity towards p-nitrophenyl butyrate at pH 8. 5 and 60°C with a Km of 0. 37 mM and a kcat of 6. 42 s(-1). It retained over 90% of its original activity after incubation at 50°C for 12 h. In addition, LipR was activated by Ca(2+), Mg(2+), Ba(2+), and Sr(2+), while strongly inhibited by Cu(2+), Zn(2+), Mn(2+), and ethylenediaminetetraacetic acid. Moreover, it showed a certain tolerance to organic solvents, including acetonitrile, isopropanol, acetone, methanol, and tert-butanol. When algal oil was hydrolyzed by LipR for 24 h, there was an enrichment of n-3 long-chain polyunsaturated fatty acids, including eicosapentaenoic acid (1. 22%, 1. 65-fold), docosapentaenoic acid (21. 24%, 2. 04-fold), and docosahexaenoic acid (36. 98%, 1. 33-fold), and even a certain amount of diacylglycerols was also produced. As a result, LipR has great prospect in industrial applications, especially in food and/or cosmetics applications.