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3 Biotech

, 9:354 | Cite as

High-level expression of lipase from Galactomyces geotrichum mafic-0601 by codon optimization in Pichia pastoris and its application in hydrolysis of various oils

  • Jian Wang
  • Yajing Liu
  • Xiaojing Guo
  • Bing Dong
  • Yunhe CaoEmail author
Original Article

Abstract

A Galactomyces geotrichum strain with lipolytic activity was isolated and identified by the analysis of internal transcribed spacer (ITS) sequence of 18 s rDNA. Full-length lipase gene of this stain is composed of 1692 base pairs (bp) without intron, which encodes a 563-amino-acid protein. A catalytic triad (Ser217–Glu354–His463) was found by constructing the three-dimensional structure of the lipase. In shake flasks, the lipase (LIP) catalytic activity in the supernatant of the recombinant Pichia pastoris increased 48.7% by codon optimization. LIP purified by anion exchange column showed a single protein band on 12% SDS-PAGE. The molecular weight (MW) of LIP was approximately 62 kDa. The specific activity of purified LIP reached 1257.9 U/mg. The optimum temperature and pH of LIP catalysis were 45 °C and pH 8.2, respectively. LIP was stable over the pH range of 4.2–11.2. LIP maintained its activity constantly at 40 °C and 50 °C for 120 min. Zn2+ inhibited LIP activity; Ba2+, Mn2+, Ca2+ and EDTA increased the enzyme activity. Referring the amount of hydrolyzed olive oil by LIP as 100%, various oils including lard, peanut oil, rapeseed oil, sunflower oil, soybean oil and linseed oil were efficiently hydrolyzed by 17.24 ± 1.34%, 40.34 ± 2.56%, 105.86 ± 2.78%, 115.51 ± 2.32%, 116.21 ± 2.15%, 120.69 ± 1.98%, respectively. The characteristics allow LIP as a potential biocatalyst in various fields of industry.

Keywords

Galactomyces geotrichum Lipase Pichia pastoris Codon optimization Characterization 

Notes

Acknowledgements

The authors sincerely thank Dr. Zhen Li for providing help on protein identification. This work was supported by National Science and Technology Support Program (2013BAD10B01) and the National High Technology Research and Development Program (2012AA022208).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

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Copyright information

© King Abdulaziz City for Science and Technology 2019

Authors and Affiliations

  1. 1.National Key Laboratory of Animal Nutrition, College of Animal Science and TechnologyChina Agricultural UniversityBeijingPeople’s Republic of China

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