Biotechnology and Bioprocess Engineering:BBE

The Korean Society for Biotechnology and Bioengineering (한국생물공학회)
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Stability Analysis of Bacillus stearothermopilus L1 Lipase Fused with a Cellulose-binding Domain

  • Hwang Sangpill (Department of Chemical Engineering, Yonsei University) ;
  • Ahn Ik-Sung (Department of Chemical Engineering, Yonsei University)
  • Published : 2005.08.01
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Abstract

This study was designed to investigate the stability of a lipase fused with a cellulose­binding domain (CBD) to cellulase. The fusion protein was derived from a gene cluster of a CBD fragment of a cellulase gene in Trichoderma hazianum and a lipase gene in Bacillus stearother­mophilus L1. Due to the CBD, this lipase can be immobilized to a cellulose material. Factors affecting the lipase stability were divided into the reaction-independent factors (RIF), and the re­action-dependent factors (RDF). RIF includes the reaction conditions such as pH and tempera­ture, whereas substrate limitation and product inhibition are examples of RDF. As pH 10 and $50^{\circ}C$ were found to be optimum reaction conditions for oil hydrolysis by this lipase, the stability of the free and the immobilized lipase was studied under these conditions. Avicel (microcrystal­line cellulose) was used as a support for lipase immobilization. The effects of both RIF and RDF on the enzyme activity were less for the immobilized lipase than for the free lipase. Due to the irreversible binding of CBD to Avicel and the high stability of the immobilized lipase, the enzyme activity after five times of use was over $70\%$ of the initial activity.

Keywords

References

  1. Park, S.-C., W.-J. Chang, S.-M. Lee, Y.-J. Kim, and Y.-M. Koo (2005) Lipase catalyzed transesterification in several reaction systems: An application of room temperature lonic liquids for bi-phasic production of n-butyl acetate, Biotechnol. Bioprocess Eng. 10: 99-102 https://doi.org/10.1007/BF02931190
  2. Pandley, A., S. Benjamin, C. R. Soccol, P. Nigam, N. Krieger, and V. T. Soccol (1999) The real of microbial lipases in biotechnology. Biotechnol. Appl. Biochem. 29: 119-131
  3. Langrand, G., N. Rondot, C. Triantaphylides, and J. C. Baratti (1990) Short chain flavour esters synthesis by microbial lipases. Biotechnol. Lett. 12: 581-586 https://doi.org/10.1007/BF01030756
  4. Talon, R., M. C. Montel, and J. L. Berdague (1996) Production of flavor esters by lipases of Staphylococcus warneri and Staphylococcus xylosus. Enzyme Microb. Technol. 19: 620-622 https://doi.org/10.1016/S0141-0229(96)00075-0
  5. Tsai, S. W., B. Y. Liu, and C. S. Chang (1996) Enhancement of (S)-naproxen ester productivity from racemic naproxen by lipase in organic solvents. J. Chem. Technol. Biotechnol. 65: 156-162 https://doi.org/10.1002/(SICI)1097-4660(199602)65:2<156::AID-JCTB415>3.0.CO;2-F
  6. Garcia, H. S., F. X. Malcata, C. G. Hill, Jr., and C. H. Amundson (1992) Use of Candida rugosa lipase immobilized in a spiral wound membrane reactor for the hydrolysis of milkfat. Enzyme Microb. Technol. 14: 535-545 https://doi.org/10.1016/0141-0229(92)90124-7
  7. Mojovic, L., S. Siler-Marinkovic, G. Kukic, and G. Vunjak-Novakovic (1993) Rhizopus arrhizus lipase-catalyzed interesterification of the midfraction of palm oil to a cocoa butter equivalent fat. Enzyme Microb. Technol. 15: 438- 443 https://doi.org/10.1016/0141-0229(93)90132-L
  8. Virto, M. D., I. Agud, S. Montero, A. Blanco, R. Solozabal, J. M. Lascaray, M. J. Llama, J. L. Serra, L. C. Landeta, and M. de Renobales (1994) Hydrolysis of animal fats by immobilized Candida rugosa lipase. Enzyme Microb. Technol. 16: 61-65 https://doi.org/10.1016/0141-0229(94)90110-4
  9. Gupta, M. N. (1991) Themostabilization of proteins. Biotechnol. Appl. Biochem. 14: 1-11
  10. Bruins, M. E., A. E. M. Janssen, and R. M. Boom (2001) Thermozymes and their applications: A review of recent literature and patents. Appl. Biochem. Biotechnol. 90: 155-186 https://doi.org/10.1385/ABAB:90:2:155
  11. Ulbrich, R., A. Schellenberger, and W. Damerau (1986) Studies on the thermal inactivation of immobilized enzymes. Biotechnol. Bioeng. 28: 511-522 https://doi.org/10.1002/bit.260280407
  12. Van der Padt, A., J. J. W. Sewalt, S. M. I. Agoston, and K. van't Riet (1992) Candida rugosa lipase stability during acylglycerol synthesis. Enzyme Microb. Technol. 14: 805-812 https://doi.org/10.1016/0141-0229(92)90096-7
  13. Ahn, J., E. Choi, H. Lee, S. Hwang, C. Kim, H. Jang, S. Haam, and J. Jung (2004) Enhanced secretion of Bacillus stearothermophilus L1 lipase in Saccharomyces cerevisiae by translational fusion to cellulose-binding domain. Appl. Microbiol. Biotechnol. 64: 833-839 https://doi.org/10.1007/s00253-003-1547-5
  14. Hwang, S., J. Ahn, S. Lee, T. G. Lee, S. Haam, K. Lee, I.- S. Ahn, and J. Jung (2004) Evaluation of cellulose-binding domain fused to a lipase for the lipase immobilization. Biotechnol. Lett. 26: 603-605 https://doi.org/10.1023/B:BILE.0000021964.69500.6f
  15. Bailey, J. E. and D. F. Ollis (1986) Biochemical Engineering Fundamentals. 2nd ed., pp. 86-156. McGraw-Hill, NY, USA
  16. Brockerhoff, H. (1968) Substrate specificity of pancreatic lipase. Biochim. Biophys. Acta. 159: 296-303 https://doi.org/10.1016/0005-2744(68)90078-8
  17. Seo, W.-Y. and K. Lee (2004) Optimized conditions for in situ immobilization of lipase in aldehyde-silica packed columns. Biotechnol. Bioprocess Eng. 9: 465-470 https://doi.org/10.1007/BF02933487
  18. Wang, T.-H. and W.-C. Lee (2003) Immobilization of proteins on magnetic nanoparticles. Biotechnol. Bioprocess Eng. 8: 263-267 https://doi.org/10.1007/BF02942276