Expression, Purification, and Characterization of a Cold-adapted Lipase from Janthinobacterium sp.
2018
Park, S.H., Chonnam National University, Gwangju, Republic of Korea | Park, S.J., Chonnam National University, Gwangju, Republic of Korea | Choi, J.I., Chonnam National University, Gwangju, Republic of Korea
The expression, purification, and characterization of cold-adapted lipase from the psychrophile, Janthinobacterium sp. were investigated. The gene encoding lipase from Janthinobacterium sp. PAMC 25641 was cloned into a pET28a(+) vector and heterologously expressed in Escherichia coli BL21 (DE3). The amino acid sequence deduced from the nucleotide sequence (930 bp) corresponded to a protein having 309 amino acid residues with a molecular weight of 32.7 kDa and a pI of 5.55. Recombinant E. coli harboring the Janthinobacterium lipase gene were induced by addition of isopropyl-beta-D-thiogalactopyranoside. Ni²+-NTA affinity chromatography was used to purify the lipase, which had a specific activity of 107.9 U/mg protein. The effect of temperature and pH on the activity of lipase was measured using p-nitrophenyl octanoate as a substrate. The stability of the lipase at low temperatures indicated it is a cold-adapted enzyme. The lipase activity was increased by Na²+, Mg²+, and Mn²+, and decreased by Zn²+ and Co²+. Analysis of the lipase activity using various p-nitrophenyl esters showed a strong preference toward short acyl chains of the esters, indicating the ability of the cold-adapted lipase to hydrolyze short-chain esters.
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