In:
Applied and Environmental Microbiology, American Society for Microbiology, Vol. 64, No. 2 ( 1998-02), p. 486-491
Abstract:
A psychrotrophic bacterium producing a cold-adapted lipase upon growth at low temperatures was isolated from Alaskan soil and identified as a Pseudomonas strain. The lipase gene ( lipP ) was cloned from the strain and sequenced. The amino acid sequence deduced from the nucleotide sequence of the gene (924 bp) corresponded to a protein of 308 amino acid residues with a molecular weight of 33,714. LipP also has consensus motifs conserved in other cold-adapted lipases, i.e., Lipase 2 from Antarctic Moraxella TA144 (G. Feller, M. Thiry, J. L. Arpigny, and C. Gerday, DNA Cell Biol. 10:381–388, 1991) and the mammalian hormone-sensitive lipase (D. Langin, H. Laurell, L. S. Holst, P. Belfrage, and C. Holm, Proc. Natl. Acad. Sci. USA 90:4897–4901, 1993): a pentapeptide, GDSAG, containing the putative active-site serine and an HG dipeptide. LipP was purified from an extract of recombinant Escherichia coli C600 cells harboring a plasmid coding for the lipP gene. The enzyme showed a 1,3-positional specificity toward triolein. p -Nitrophenyl esters of fatty acids with short to medium chains (C 4 and C 6 ) served as good substrates. The enzyme was stable between pH 6 and 9, and the optimal pH for the enzymatic hydrolysis of tributyrin was around 8. The activation energies for the hydrolysis of p -nitrophenyl butyrate and p -nitrophenyl laurate were determined to be 11.2 and 7.7 kcal/mol, respectively, in the temperature range 5 to 35°C. The enzyme was unstable at temperatures higher than 45°C. The K m of the enzyme for p -nitrophenyl butyrate increased with increases in the assay temperature. The enzyme was strongly inhibited by Zn 2+ , Cu 2+ , Fe 3+ , and Hg 2+ but was not affected by phenylmethylsulfonyl fluoride and bis-nitrophenyl phosphate. Various water-miscible organic solvents, such as methanol and dimethyl sulfoxide, at concentrations of 0 to 30% (vol/vol) activated the enzyme.
Type of Medium:
Online Resource
ISSN:
0099-2240
,
1098-5336
DOI:
10.1128/AEM.64.2.486-491.1998
Language:
English
Publisher:
American Society for Microbiology
Publication Date:
1998
detail.hit.zdb_id:
223011-2
detail.hit.zdb_id:
1478346-0
SSG:
12
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