Cloning, Expression, and Characterization of a Cold-Adapted Lipase Gene from an Antarctic Deep-Sea Psychrotrophic Bacterium, Psychrobacter sp. 7195
A psychrotrophic strain 7195 showing extracellular lipolytic activity towards tributyrin was isolated from deep-sea sediment of Prydz Bay and identified as a Psychrobacter species. By screening a genomic DNA library of Psychrobacter sp. 7195, an open reading frame of 954 bp coding for a lipase gene,...
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The Korean Society for Applied Microbiology and Biotechnology
2008
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ftstoai:JAKO200717317773780 2023-05-15T13:44:16+02:00 Cloning, Expression, and Characterization of a Cold-Adapted Lipase Gene from an Antarctic Deep-Sea Psychrotrophic Bacterium, Psychrobacter sp. 7195 Zhang, Jinwei Lin, Shu Zeng, Runying 2008-01-29 pdf http://click.ndsl.kr/servlet/LinkingFullTextView?service_code=04&dbt=JAKO&cn=JAKO200717317773780 en eng The Korean Society for Applied Microbiology and Biotechnology 66062 2008 ftstoai 2009-03-30T20:34:36Z A psychrotrophic strain 7195 showing extracellular lipolytic activity towards tributyrin was isolated from deep-sea sediment of Prydz Bay and identified as a Psychrobacter species. By screening a genomic DNA library of Psychrobacter sp. 7195, an open reading frame of 954 bp coding for a lipase gene, lipA1, was identified, cloned, and sequenced. The deduced LipA1 consisted of 317 amino acids with a molecular mass of 35,210 kDa. It had one consensus motif, G-N-S-M-G (GXSXG), containing the putative active-site serine, which was conserved in other cold-adapted lipolytic enzymes. The recombinant LipA1 was purified by column chromatography with DEAE Sepharose CL-4B, and Sephadex G-75, and preparative polyacrylamide gel electrophoresis, in sequence. The purified enzyme showed highest activity at $30^{\circ}C$ , and was unstable at temperatures higher than $30^{\circ}C$ , indicating that it was a typical cold-adapted enzyme. The optimal pH for activity was 9.0, and the enzyme was stable between pH 7.0-10.0 after 24h incubation at $4^{\circ}C$ . The addition of $Ca^{2+}\;and\;Mg^{2+}$ enhanced the enzyme activity of LipA1, whereas the $Cd^{2+},\;Zn^{2+},\;CO^{2+},\;Fe^{3+},\;Hg^{2+},\;Fe^{2+},\;Rb^{2+}$ , and EDTA strongly inhibited the activity. The LipA1 was activated by various detergents, such as Triton X-100, Tween 80, Tween 40, Span 60, Span 40, CHAPS, and SDS, and showed better resistance towards them. Substrate specificity analysis showed that there was a preference for trimyristin and p-nitrophenyl myristate $(C_{14}\;acyl\; groups)$ . Other/Unknown Material Antarc* Antarctic Prydz Bay CLICK (Cooperative Link Center in KOREA) Antarctic Prydz Bay Triton ENVELOPE(-55.615,-55.615,49.517,49.517) |
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CLICK (Cooperative Link Center in KOREA) |
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language |
English |
topic |
66062 |
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66062 Zhang, Jinwei Lin, Shu Zeng, Runying Cloning, Expression, and Characterization of a Cold-Adapted Lipase Gene from an Antarctic Deep-Sea Psychrotrophic Bacterium, Psychrobacter sp. 7195 |
topic_facet |
66062 |
description |
A psychrotrophic strain 7195 showing extracellular lipolytic activity towards tributyrin was isolated from deep-sea sediment of Prydz Bay and identified as a Psychrobacter species. By screening a genomic DNA library of Psychrobacter sp. 7195, an open reading frame of 954 bp coding for a lipase gene, lipA1, was identified, cloned, and sequenced. The deduced LipA1 consisted of 317 amino acids with a molecular mass of 35,210 kDa. It had one consensus motif, G-N-S-M-G (GXSXG), containing the putative active-site serine, which was conserved in other cold-adapted lipolytic enzymes. The recombinant LipA1 was purified by column chromatography with DEAE Sepharose CL-4B, and Sephadex G-75, and preparative polyacrylamide gel electrophoresis, in sequence. The purified enzyme showed highest activity at $30^{\circ}C$ , and was unstable at temperatures higher than $30^{\circ}C$ , indicating that it was a typical cold-adapted enzyme. The optimal pH for activity was 9.0, and the enzyme was stable between pH 7.0-10.0 after 24h incubation at $4^{\circ}C$ . The addition of $Ca^{2+}\;and\;Mg^{2+}$ enhanced the enzyme activity of LipA1, whereas the $Cd^{2+},\;Zn^{2+},\;CO^{2+},\;Fe^{3+},\;Hg^{2+},\;Fe^{2+},\;Rb^{2+}$ , and EDTA strongly inhibited the activity. The LipA1 was activated by various detergents, such as Triton X-100, Tween 80, Tween 40, Span 60, Span 40, CHAPS, and SDS, and showed better resistance towards them. Substrate specificity analysis showed that there was a preference for trimyristin and p-nitrophenyl myristate $(C_{14}\;acyl\; groups)$ . |
author |
Zhang, Jinwei Lin, Shu Zeng, Runying |
author_facet |
Zhang, Jinwei Lin, Shu Zeng, Runying |
author_sort |
Zhang, Jinwei |
title |
Cloning, Expression, and Characterization of a Cold-Adapted Lipase Gene from an Antarctic Deep-Sea Psychrotrophic Bacterium, Psychrobacter sp. 7195 |
title_short |
Cloning, Expression, and Characterization of a Cold-Adapted Lipase Gene from an Antarctic Deep-Sea Psychrotrophic Bacterium, Psychrobacter sp. 7195 |
title_full |
Cloning, Expression, and Characterization of a Cold-Adapted Lipase Gene from an Antarctic Deep-Sea Psychrotrophic Bacterium, Psychrobacter sp. 7195 |
title_fullStr |
Cloning, Expression, and Characterization of a Cold-Adapted Lipase Gene from an Antarctic Deep-Sea Psychrotrophic Bacterium, Psychrobacter sp. 7195 |
title_full_unstemmed |
Cloning, Expression, and Characterization of a Cold-Adapted Lipase Gene from an Antarctic Deep-Sea Psychrotrophic Bacterium, Psychrobacter sp. 7195 |
title_sort |
cloning, expression, and characterization of a cold-adapted lipase gene from an antarctic deep-sea psychrotrophic bacterium, psychrobacter sp. 7195 |
publisher |
The Korean Society for Applied Microbiology and Biotechnology |
publishDate |
2008 |
url |
http://click.ndsl.kr/servlet/LinkingFullTextView?service_code=04&dbt=JAKO&cn=JAKO200717317773780 |
long_lat |
ENVELOPE(-55.615,-55.615,49.517,49.517) |
geographic |
Antarctic Prydz Bay Triton |
geographic_facet |
Antarctic Prydz Bay Triton |
genre |
Antarc* Antarctic Prydz Bay |
genre_facet |
Antarc* Antarctic Prydz Bay |
_version_ |
1766199548675358720 |