High-level heterologous expression and properties of a novel lipase from Ralstonia sp. M1

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dc.contributor.authorD T Quyen-
dc.contributor.authorT T G Le-
dc.contributor.authorT T Nguyen-
dc.contributor.authorTae Kwang Oh-
dc.contributor.authorJung-Kee Lee-
dc.date.accessioned2017-04-19T09:02:05Z-
dc.date.available2017-04-19T09:02:05Z-
dc.date.issued2005-
dc.identifier.issn1046-5928-
dc.identifier.uri10.1016/j.pep.2004.10.001ko
dc.identifier.urihttps://oak.kribb.re.kr/handle/201005/6780-
dc.description.abstractThe mature lipase LipA and its 56aa-truncated chaperone ΔLipBhis (with 6×his-tag) from Ralstonia sp. M1 were over-expressed in Escherichia coli BL21 under the control of T7 promoter with a high level of 70 and 12 mg protein per gram of wet cells, respectively. The simply purified lipase LipA was effectively refolded by Ni-NTA purified chaperone ΔLipBhis in molar ratio 1:1 at 4°C for 24 hours in H2O. The in vitro refolded lipase LipA had an optimal activity in the temperature range of 50-55°C and was stable up to 45°C with more than 84% activity retention. The maximal activity was observed at pH 10.75 for hydrolysis of olive oil and found to be stable over alkaline pH range 8.0-10.5 with more than 52% activity retention. The enzyme was found to be highly resistant to many organic solvents especially induced by ethanolamine (remaining activity 137-334%), but inhibited by 1-butanol and acetonitrile (40-86%). Metal ions Cu2+, Sn 2+, Mn2+, Mg2+, and Ca2+ stimulated the lipase slightly with increase in activity by up to 22%, whereas Zn 2+ significantly inhibited the enzyme with the residual activity of 30-65% and Fe3+ to a lesser degree (activity retention of 77-86%). Tween 80, Tween 60, and Tween 40 induced the activation of the lipase LipA (222-330%) and 0.2-1% (w/v) of Triton X-100, X-45, and SDS increased the lipase activity by up to 52%. However, 5% (w/v) of Triton X-100, X-45, and SDS inhibited strongly the activity by 31-89%. The inhibitors including DEPC, EDTA, PMSF, and 2-mercaptoethanol (0.1-10 mM) inhibited moderately the lipase with remaining activity of 57-105%. The lipase LipA hydrolyzed a wide range of triglycerides, but preferentially short length acyl chains (C4 and C6). In contrast to the triglycerides, medium length acyl chains (C8 and C14) of p-nitrophenyl (p-NP) esters were preferential substrates of this lipase. The enzyme preferentially catalyzed the hydrolysis of cottonseed oil (317%), cornoil (227%), palm oil (222%), and wheatgerm oil (210%) in comparison to olive oil (100%).-
dc.publisherElsevier-
dc.titleHigh-level heterologous expression and properties of a novel lipase from Ralstonia sp. M1-
dc.title.alternativeHigh-level heterologous expression and properties of a novel lipase from Ralstonia sp. M1-
dc.typeArticle-
dc.citation.titleProtein Expression and Purification-
dc.citation.number1-
dc.citation.endPage106-
dc.citation.startPage97-
dc.citation.volume39-
dc.contributor.affiliatedAuthorTae Kwang Oh-
dc.contributor.affiliatedAuthorJung-Kee Lee-
dc.contributor.alternativeNameQuyen-
dc.contributor.alternativeNameLe-
dc.contributor.alternativeNameNguyen-
dc.contributor.alternativeName오태광-
dc.contributor.alternativeName이정기-
dc.identifier.bibliographicCitationProtein Expression and Purification, vol. 39, no. 1, pp. 97-106-
dc.identifier.doi10.1016/j.pep.2004.10.001-
dc.subject.keywordChaperone-
dc.subject.keywordE. coli-
dc.subject.keywordLipase-
dc.subject.keywordOver-expression-
dc.subject.keywordProperties-
dc.subject.keywordRalstonia sp. M1-
dc.subject.localchaperone-
dc.subject.localChaperone-
dc.subject.localEscherichia coli.-
dc.subject.localescherichia coli-
dc.subject.localEscherichia Coli-
dc.subject.localEscherichia coli-
dc.subject.localE.coli-
dc.subject.localescherichia coil-
dc.subject.localE. coli-
dc.subject.localE. Coli-
dc.subject.locallipase-
dc.subject.localLipase-
dc.subject.localOverexpression-
dc.subject.localoverexpression-
dc.subject.localover-expression-
dc.subject.localOver-expression-
dc.subject.localproperty-
dc.subject.localProperties-
dc.subject.localProperty-
dc.subject.localproperties-
dc.subject.localRalstonia sp. M1-
dc.description.journalClassY-
Appears in Collections:
Division of Biomedical Research > Metabolic Regulation Research Center > 1. Journal Articles
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