Co-expression, purification and characterization of the lipase and foldase of Burkholderia contaminans LTEB11

被引：15

作者：

Carlos Alnoch, Robson ^{[1
,2
]}

Stefanello, Adriano Alves ^{[1
]}

Martini, Viviane Paula ^{[3
]}

Richter, Jeferson Luiz ^{[1
]}

Mateo, Cesar ^{[2
]}

de Souza, Emanuel Maltempi ^{[1
]}

Mitchell, David Alexander ^{[1
]}

Muller-Santos, Marcelo ^{[1
]}

Krieger, Nadia ^{[4
]}

机构：

[1] Univ Fed Parana, Ctr Politecn, Dept Bioquim & Biol Mol, Cx P 19046, BR-81531980 Curitiba, Parana, Brazil

[2] Inst Catalisis & Petr Quim CSIC, Dept Biocatalisis, Marie Curie 2,Campus UAM, Madrid 28049, Spain

[3] Inst Fed Parana, Campus Irati, BR-84500000 Irati, Parana, Brazil

[4] Univ Fed Parana, Dept Quim, Ctr Politecn, Cx P 19081, BR-81531980 Curitiba, Parana, Brazil

来源：

INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES | 2018年 / 116卷

关键词：

Lipases; Foldases; Burkholderia contaminans; Co-expression in vivo; Ester synthesis; EXTRACELLULAR LIPASE; SEQUENCE ALIGNMENT; BACTERIAL LIPASES; PSEUDOMONAS; CHAPERONE; GENE; EXPRESSION; BIODIESEL; IMMOBILIZATION; IDENTIFICATION;

D O I：

10.1016/j.ijbiomac.2018.05.086

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

Genes encoding lipase LipBC (lipA) and foldase LifBC (lipB) were identified in the genome of Burkholderia contaminans LTEB11. Analysis of the predicted amino acid sequence of lipA showed its high identity with lipases from Pseudomonas luteola (91%), Burkholderia cepacia (96%) and Burkholderia Iota (97%), and classified LipBC lipase in the lipase subfamily 1.2. The genes lipA and lipB were amplified and cloned into expression vectors pET28a(+) and pT7-7, respectively. His-tagged LipBC and native LifBC were co-expressed in Escherichia coli and purified. LipBC and LifBC have molecular weights of 35.9 kDa and 37 kDa, respectively, and remain cornplexed after purification. The Lip-LifBC complex was active and stable over a wide range of pH values (6.5-10) and temperatures (25-45 degrees C), with the highest specific activity (1426 U mg(-1)) being against tributyrin. The Lip-LifBC complex immobilized on Sepabeads was able to catalyze the synthesis of ethyl-oleate in n hexane with an activity of 4 U g(-1), maintaining high conversion (>80%) over 5 reaction cycles of 6 hat 45 degrees C. The results obtained in this work provide a basis for the development of applications of recombinant LipBC in biocatalysis. (C) 2018 Elsevier B.V. All rights reserved.

引用

页码：1222 / 1231

页数：10

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