Skip to main content
SpringerLink
Log in

Gene Cloning and Characterization of a Novel Highly Organic Solvent Tolerant Lipase from Proteus sp. SW1 and its Application for Biodiesel Production

  • Research
  • Published:
Molecular Biotechnology Aims and scope Submit manuscript

Abstract

Proteus sp. SW1 was found to produce an extracellular solvent tolerant lipase. The gene, lipA, encoding a bacterial lipase, was cloned from total Proteus sp. SW1 DNA. lipA was predicted to encode a 287 amino acid protein of 31.2 kDa belonging to the Group I proteobacterial lipases. Purified His-tagged LipA exhibited optimal activity at pH 10.0 and 55°C. It was highly stable in organic solvents retaining 112% of its activity in 100% isopropanol after 24 h, and exhibited more than 200% of its initial activity upon exposure to 60% acetone, ethanol, and hexane for 18 h. Biodiesel synthesis reactions, using a single step addition of 13% an acyl acceptor ethanol, showed that LipA was highly effective at converting palm oil into biodiesel.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. Cho, S. S., Park, D. J., Simkhada, J. R., Hong, J. H., Sohng, J. K., Lee, O. H., et al. (2012). A neutral lipase applicable in biodiesel production from a newly isolated Streptomyces sp. CS326. Bioprocess and Biosystems Engineering, 35, 227–234.

    Article  CAS  Google Scholar 

  2. Simkhada, J. R., Yoo, H. Y., Cho, S. S., Choi, Y. H., Kim, S. W., Park, D. H., et al. (2012). A novel cold-adapted lipase, LP28, from a mesophilic Streptomyces strain. Bioprocess and Biosystems Engineering, 35, 217–225.

    Article  CAS  Google Scholar 

  3. Li, Z., Li, X., Wang, Y., Wang, F., & Jiang, J. (2011). Expression and characterization of recombinant rhizopus oryzae lipase for enzymatic biodiesel production. Bioresource Technology, 102, 9810–9813.

    Article  CAS  Google Scholar 

  4. Yoo, H. Y., Simkhada, J. R., Cho, S. S., Park, D. H., Kim, S. W., Seong, C. N., et al. (2011). A novel alkaline lipase from ralstonia with potential application in biodiesel production. Bioresource Technology, 102, 6104–6111.

    Article  CAS  Google Scholar 

  5. Parawira, W. (2009). Biotechnological production of biodiesel fuel using biocatalysed transesterification: a review. Critical Review in Biotechnology, 29, 82–93.

    Article  CAS  Google Scholar 

  6. Sulong, M. R., Abdul Rahman, R. N., Salleh, A. B., & Basri, M. (2006). A novel organic solvent tolerant lipase from Bacillus sphaericus 205y: extracellular expression of a novel OST-lipase gene. Protein Expression and Purification, 49, 190–195.

    Article  CAS  Google Scholar 

  7. Madan, B., & Mishra, P. (2009). Overexpression, purification and characterization of organic solvent stable lipase from Bacillus licheniformis RSP-09. Journal of Molecular Microbiology and Biotechnology, 17, 118–123.

    Article  CAS  Google Scholar 

  8. Ogino, H., Katou, Y., Akagi, R., Mimitsuka, T., Hiroshima, S., Gemba, Y., et al. (2007). Cloning and expression of gene, and activation of an organic solvent-stable lipase from Pseudomonas aeruginosa LST-03. Extremophiles, 11, 809–817.

    Article  CAS  Google Scholar 

  9. Arpigny, J. L., & Jaeger, K. E. (1999). Bacterial lipolytic enzymes: classification and properties. Biochemistry Journal, 343, 177–183.

    Article  CAS  Google Scholar 

  10. Gilham, D., & Lehner, R. (2005). Techniques to measure lipase and esterase activity in vitro. Methods, 36, 139–147.

    Article  CAS  Google Scholar 

  11. Yang, K. S., Sohn, J. H., & Kim, H. K. (2009). Catalytic properties of a lipase from Photobacterium lipolyticum for biodiesel production containing a high methanol concentration. Journal of Bioscience and Bioengineering, 107, 599–604.

    Article  CAS  Google Scholar 

  12. Winayanuwattikun, P., Kaewpiboon, C., Piriyakananon, K., Tantong, S., Thakernkarnkit, W., Chulalaksananukul, W., et al. (2008). Potential plant oil feedstock for lipase-catalyzed biodiesel production in Thailand. Biomass and Bioenergy, 32, 1279–1286.

    Article  CAS  Google Scholar 

  13. Sullivan, E. R., Leahy, J. G., & Colwell, R. R. (1999). Cloning and sequence analysis of the lipase and lipase chaperone-encoding genes from Acinetobacter calcoaceticus RAG-1, and redefinition of a proteobacterial lipase family and an analogous lipase chaperone family. Gene, 230, 277–286.

    Article  CAS  Google Scholar 

  14. Kim, H. K., Lee, J. K., Kim, H., & Oh, T. K. (1996). Characterization of an alkaline lipase from Proteus vulgaris K80 and the DNA sequence of the encoding gene. FEMS Microbiology Letters, 135, 117–121.

    Article  CAS  Google Scholar 

  15. Brumlik, M. J., & Buckley, J. T. (1996). Identification of the catalytic triad of the lipase/acyltransferase from Aeromonas hydrophila. Journal of Bacteriology, 178, 2060–2064.

    CAS  Google Scholar 

  16. Jaeger, K. E., Ransac, S., Dijkstra, B. W., Colson, C., van Heuvel, M., & Misset, O. (1994). Bacterial lipases. FEMS Microbiology Reviews, 15, 29–63.

    Article  CAS  Google Scholar 

  17. Ahmed, E. H., Raghavendra, T., & Madamwar, D. (2010). An alkaline lipase from organic solvent tolerant Acinetobacter sp. EH28: application for ethyl caprylate synthesis. Bioresouce Technology, 101, 3628–3634.

    Article  CAS  Google Scholar 

  18. Karadzic, I., Masui, A., Zivkovic, L. I., & Fujiwara, N. (2006). Purification and characterization of an alkaline lipase from Pseudomonas aeruginosa isolated from putrid mineral cutting oil as component of metalworking fluid. Journal of Bioscience and Bioengineering, 102, 82–89.

    Article  CAS  Google Scholar 

  19. Snellman, E. A., Sullivan, E. R., & Colwell, R. R. (2002). Purification and properties of the extracellular lipase, LipA, of Acinetobacter sp. RAG-1. European Journal of Biochemistry, 269, 5771–5779.

    Article  CAS  Google Scholar 

  20. Salameh, M. A., & Wiegel, J. (2010). Effects of detergents on activity, thermostability and aggregation of two alkalithermophilic lipases from Thermosyntropha lipolytica. Open Biochemistry Journal, 4, 22–28.

    Article  CAS  Google Scholar 

  21. Mogensen, J. E., Sehgal, P., & Otzen, D. E. (2005). Activation, inhibition, and destabilization of Thermomyces lanuginosus lipase by detergents. Biochemistry, 44, 1719–1730.

    Article  CAS  Google Scholar 

  22. Quyen, D. T., Le Giang, T. T., Nguyen, T. T., Oh, T. K., & Lee, J. K. (2005). High-level heterologous expression and properties of a novel lipase from Ralstonia sp. M1. Protein Expression and Purification, 39, 97–106.

    Article  CAS  Google Scholar 

  23. Klibanov, A. M. (2001). Improving enzymes by using them in organic solvents. Nature, 409, 241–246.

    Article  CAS  Google Scholar 

  24. Matsumoto, M., Kida, K., & Kondo, K. (2001). Enhanced activities of lipase pretreated with organic solvents. Journal of Chemical Technology and Biotechnology, 76, 1070–1073.

    Article  CAS  Google Scholar 

  25. Ebrahimpour, A., Rahman, R. N., Basri, M., & Salleh, A. B. (2011). High level expression and characterization of a novel thermostable, organic solvent tolerant, 1,3-regioselective lipase from Geobacillus sp. strain ARM. Bioresource Technology, 102, 6972–6981.

    Article  CAS  Google Scholar 

  26. Hong, M. C., & Chang, M. C. (1998). Purification and characterization of an alkaline lipase from a newly isolated Acinetobacter radioresistens CMC-1. Biotechnology Letters, 20, 1027–1029.

    Article  CAS  Google Scholar 

  27. Samukawa, T., Kaieda, M., Matsumoto, T., Ban, K., Kondo, A., Shimada, Y., et al. (2000). Pretreatment of immobilized Candida antarctica lipase for biodiesel fuel production from plant oil. Journal of Bioscience and Bioengineering, 90, 180–183.

    CAS  Google Scholar 

Download references

Acknowledgments

We thank N. Thasana and S. Thumniyom for GC–MS analysis, P. Winayanuwattikun and K. Piriyakananon for HPLC analysis, P. Munpiyamit for the photograph preparation, and J. Dubbs for critical reading and discussion. This research was supported by grants from the Chulabhorn Research Institute and the Center of Excellence on Environmental Health and Toxicology, Science & technology Postgraduate Education and Research Development Office (PERDO), Ministry of Education, Thailand to S.L.

Author information

Authors and Affiliations

  1. Laboratory of Biotechnology, Chulabhorn Research Institute, Kamphaengphet 6 Road, Bangkok, 10210, Thailand

    Wirongrong Whangsuk, Jarunee Kerdwong, Ratiboot Sallabhan, Skorn Mongkolsuk & Suvit Loprasert

  2. Applied Biological Sciences Program, Chulabhorn Graduate Institute, Bangkok, 10210, Thailand

    Pareenart Sungkeeree, Sirinthra Thiengmag & Suvit Loprasert

  3. Center of Excellence on Environmental Health and Toxicology, CHE, Ministry of Education, Bangkok, Thailand

    Skorn Mongkolsuk & Suvit Loprasert

Authors
  1. Wirongrong Whangsuk
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Pareenart Sungkeeree
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Sirinthra Thiengmag
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jarunee Kerdwong
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Ratiboot Sallabhan
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Skorn Mongkolsuk
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Suvit Loprasert
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Suvit Loprasert.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Whangsuk, W., Sungkeeree, P., Thiengmag, S. et al. Gene Cloning and Characterization of a Novel Highly Organic Solvent Tolerant Lipase from Proteus sp. SW1 and its Application for Biodiesel Production. Mol Biotechnol 53, 55–62 (2013). https://doi.org/10.1007/s12033-012-9518-7

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12033-012-9518-7

Keywords

Search

Navigation