Article Search
닫기

Microbiology and Biotechnology Letters

보문(Article)

View PDF

Biocatalysis and Bioprocess Engineering  |  Enzyme, Protein Engineering, and Metabolic Engineering

Microbiol. Biotechnol. Lett. 2018; 46(3): 234-243

https://doi.org/10.4014/mbl.1804.04012

Received: April 12, 2018; Accepted: June 25, 2018

남극 로스해에서 분리한 Croceibacter atlanticus 균 유래 리파아제의 생산, 고정화, 효소특성 연구

Production, Immobilization, and Characterization of Croceibacter atlanticus Lipase Isolated from the Antarctic Ross Sea

Chae Gyeong Park and Hyung Kwoun Kim *

Division of Biotechnology, The Catholic University of Korea

The Antarctic Ocean contains numerous microorganisms that produce novel biocatalysts that can have applications in various industries. We screened various psychrophilic bacterial strains isolated from the Ross Sea and found that a Croceibacter atlanticus strain (Stock No. 40-F12) showed high lipolytic activity on a tributyrin plate. We isolated the corresponding lipase gene (lipCA) by shotgun cloning and expressed the LipCA enzyme in Escherichia coli cells. Homology modeling of LipCA was carried out using the Spain Arreo lake metagenome alpha/beta hydrolase as a template. According to the model, LipCA has an α/β hydrolase fold, Gly-X-Ser-X-Glymotif, and lid sequence, indicating that LipCA is a typical lipase enzyme. Active LipCA enzyme was purified fromthe cell-free extract by ammonium sulfate precipitation and gel filtration chromatography. We determined its enzymatic properties including optimum temperature and pH, stability, substrate specificity, and organic solvent stability. LipCA was immobilized by the cross-linked enzyme aggregate (CLEA) method and its enzymatic properties were compared to those of free LipCA. After cross-linking, temperature, pH, and organic solvent stability increased considerably, whereas substrate specificities did not changed. The LipCA CLEA was recovered by centrifugation and showed approximately 40% activity after 4th recovery. This is the first report of the expression, characterization, and immobilization of a C. atlanticus lipase, and this lipase could have potential industrial application.

Keywords: Lipase, Croceibacter atlanticus, the Antarctic Ross Sea

  1. Sharma R, Chisti Y, Banerjee UC. 2001. Production, purification, characterization, and applications of lipase. Biotechnol. Adv. 19:627-662.
    CrossRef
  2. Datta S, Christena LR, Rajaram YRS. 2013. Enzyme immobilization:an overview on techniques and support materials. 3 Biotech. 3: 1-9.
  3. Sheldon RA. 2011. Cross-linked enzyme aggregates as industrial biocatalysts. Org. Process Res. Dev. 15: 213-223.
    CrossRef
  4. Lenfant N, Hotelier T, Velluet E, Bourne Y, Marchot P, Chatonnet A. 2013. ESTHER, the database of the α/β-hydrolase fold superfamily of proteins: tools to explore diversity of functions. Nucl. Acids Res. 41: 423-429.
    Pubmed KoreaMed CrossRef
  5. Arpigny JL, Jaeger KE. 1999. Bacterial lipolytic enzymes: classification and properties. Biochem. J. 343: 177-183.
    Pubmed KoreaMed CrossRef
  6. Martinez-Martinez M, Alcaide M, Tchigvintsev A, Reva O, Polaina J, Bargiela R, et al. 2013. Biochemical diversity of carboxyl esterases and lipases from lake Arreo (Spain): a metagenomic approach. Appl. Environ. Microbiol. 79: 3553-3562.
    Pubmed KoreaMed CrossRef
  7. Kartal F, Janssen MHA, Hollmann F, Sheldon RA, Kilinc A. 2011. Improved esterification activity of Candida rugosa lipase in organic solvent by immobilization as cross-linked enzyme aggregates (CLEAs). J. Mol. Catal. B: Enzym. 71: 85-89.
    CrossRef
  8. Rehman S, Bhatti HN, Bilal M, Asgher M. 2016. Cross-linked enzyme aggregates (CLEAs) of Pencilluim notatum lipase enzyme with improved activity, stability and reusability characteristics. Int. J. Biol. Macromol. 91: 1161-1169.
    Pubmed CrossRef
  9. Iftikhar T, Niaz M, Jabeen R, Haq IU. 2011. Purification and characterization of extracellular lipase. Pak. J. Bot. 43: 1541-1545.
  10. Pérez D, Martín S, Fernández-Lorente G, Filice M, Guisán JM, Ventosa A, et al. 2011. A novel halophilic lipase, LipBL, showing high efficiency in the production of eicosapentaenoic acid (EPA). PLoS One. https://doi.org/10.1371/journal.pone.0023325.
    CrossRef
  11. Kim HK, Park SY, Lee JK, Oh TK. 1998. Gene cloning and characterization of thermal stable lipase from Bacillus stearothermophilus L1. Biosci. Biotechnol. Biochem. 62: 66-71.
    Pubmed CrossRef
  12. Cho JC, Giovannoni SJ. 2003. Croceibacter atlanticus gen.nov., sp. Nov., A Novel Marine Bacterium in the Family Flavobacteriaceae. Syst. Appl. Microbiol. 26: 76-83.
    Pubmed CrossRef
  13. Lai Q, Wang J, Gu L, Zheng T, Shao Z. 2013. Alcanivorax marinus sp. Nov., isolated from deep-sea water. Int. J. Syst. Evol. Microbiol. 63: 4428-4432.
    Pubmed CrossRef
  14. Saxena RK, Sheoran A, Giri B, Davidson WS. 2003. Purification strategies for microbial lipase. J. Microbiol. Methods. 52: 1-18.
    CrossRef
  15. Li M, Yang LR, Xu G, Wu JP. 2013. Screening, purification and characterization of a novel cold-active and organic solvent-tolerant lipase from Stenotrophomonas maltophilia CGMCC 4254. Bioresour. Technol. 148: 114-120.
    Pubmed CrossRef
  16. Wang Q, Hou Y, Ding Y, Yan P. 2012. Purification and biochemical characterization of a cold-active lipase from Antarctic sea ice bacteria Pseudoalteromonas sp. NJ 70. Mol. Biol. Rep. 39:9233-9238.
    Pubmed CrossRef
  17. Gauthier MA, Ayer M, Kowal J, Wurm FR, Klok HA. 2011. Argininespecific protein modification using α-oxo-aldehyde functional polymers prepared by atom transfer radical polymerization. Polym. Chem. 2: 1490-1498.
    CrossRef
  18. Farris S, Song J, Huang Q. 2010. Alternative reaction mechanism for the cross-linking of gelatin with glutaraldehyde. J. Agric. Food Chem. 58: 998-1003.
    Pubmed CrossRef
  19. Migneault I, Dartiguenave C, Bertrand MJ, Waldron KC. 2004. Glutaraldehyde: behavior in aqueous solution, reaction with proteins, and application to enzyme crosslinking. Biotechniques. 37: 790-802.
    Pubmed CrossRef
  20. Yedavalli P, Rao NM. 2013. Engineering the loops in a lipase for stability in DMSO. Protein Eng. Des. Sel. 26: 317-324.
    Pubmed CrossRef
  21. Dachuri V, Boyineni J, Choi S, Chung HS, Jang SH, Lee CW. 2016. Organic solvent-tolerant, cold-adapted lipase PML and LipS exhibit increased conformational flexibility in polar organic solvents. J. Mol. Catal. B: Enzym. 131: 73-78.
    CrossRef
  22. Lopez-Serrano P, Cao L, van Rantwijk F, Sheldon RA. 2002. Cross-linked enzyme aggregates with enhanced activity:application to lipases. Biotechnol. Lett. 24: 1379-1383.
    CrossRef
  23. Valdes EC, Soto LW, Arcaya GA. 2011. Influence of the pH of glutaraldehyde and the use of dextran aldehyde on the preparaton of cross-linked enzme aggregates (CLEAs) of lipase from Burkholderia cepacia. Electronic J. Biotechnol. 14. doi:10.2225/vol14-issue3-fulltext-1.
    CrossRef

Starts of Metrics

Share this article on :

  • mail

Related articles in MBL

Most KeyWord ?

What is Most Keyword?

  • It is most registrated keyword in articles at this journal during for 2 years.