Abstract

Biodiesel has attracted considerable attention as an environmentally friendly alternative fuel. Lipase is the most popular enzyme for biodiesel production and immobilization has been deployed to improve enzyme stability and reusability. Exploitation of high activity lipase is the key point for biodiesel production. Whole-cell biocatalysts have been applied in the biosynthesis of biodiesel and bioimprinting is a promising approach for enzyme performances improvement. In this study, based on the S. cerevisiae cell-surface display system with $\alpha$-agglutinin as anchor, a whole-cell biocatalyst of codon-optimized Rhizopus oryzae lipase was constructed and bioimprinted with oleic acid, gaining 5-fold increase on enzymatic activity in the alcoholysis of soybean oil to biodiesel. Moreover, the conversion of FAME was up to 95.45±2.73% after a 27-h reaction at 60°C. Our results indicated that combining bioimprinting with yeast display technique to prepare whole-cell biocatalyst could result in potential enzymes for bioconversion of biodiesel in organic solvents.

Keywords:

Biodiesel, bioimprinting, Rhizopus oryzae Lipase (ROL), whole-cell biocatalyst, yeast surface display,

References

1. Adamczak, M., U.T. Bornscheuer and W. Bednarski, 2009. The application of biotechnological methods for the synthesis of biodiesel. Eur. J. Lipid Sci. Tech., 111(8): 800-813.
   CrossRef    
2. Arai, S., K. Nakashima, T. Tanino, C. Ogino, A. Kondo and H. Fukuda, 2010. Production of biodiesel fuel from soybean oil catalyzed by fungus whole-cell biocatalysts in ionic liquids. Enzyme Microb. Tech., 46(1): 51-55.
   CrossRef    Direct Link
3. Caballero, V., F.M. Bautista, J.M. Campelo, D. Luna, J.M. Marinas, A.A. Romero, J.M. Hidalgo, R. Luque, A. Macario and G. Giordano, 2009. Sustainable preparation of a novel glycerol-free biofuel by using pig pancreatic lipase: Partial 1,3-regiospecific alcoholysis of sunflower oil. Process Biochem., 44(3): 334-342.
   CrossRef    Direct Link
4. Chen, M.L., Y. He, G.Q. He and H. Ruan, 2012. The preparation of a hyper-thermostable whole-cell biocatalyst and its application for biosynthesis of biodiesel. Adv. Mater. Res., 550-553: 1381-1386.
   CrossRef    
5. Chen, M.L., Q. Guo, R.Z. Wang, J. Xu, C.W. Zhou, H. Ruan and G.Q. He, 2011. Construction of the yeast whole-cell Rhizopus oryzae lipase biocatalyst with high activity. J. Zhejiang Univ. Sci. B., 12(7): 545-551.
   CrossRef    PMid:21726061 PMCid:PMC3134842    
6. Fishman, A. and U. Cogan, 2003. Bio-imprinting of lipases with fatty acids. J. Mol. Catal. B-Enzym., 22(3-4): 193-202.
   CrossRef    Direct Link
7. Fukuda, H., S. Hama, S. Tamalampudi and H. Noda, 2008. Whole-cell biocatalysts for biodiesel fuel production. Trends Biotechnol., 26(12): 668-673.
   CrossRef    PMid:18976825    
8. Gonzalez-Navarro, H. and L. Braco, 1997. Improving lipase activity in solvent-free media by interfacial activation-based molecular bioimprinting. J. Mol. Catal. B-Enzym., 3(1-4): 111-119.
   CrossRef    
9. Gonzalez-Navarro, H. and L. Braco, 1998. Lipase-enhanced activity in flavour ester reactions by trapping enzyme conformers in the presence of interfaces. Biotechnol. Bioeng., 59(1): 122-127.
   CrossRef    
10. Hama, S., S. Tamalampudi, T. Fukumizu, K. Miura, H. Yamaji, A. Kondo and H. Fukuda, 2006. Lipase localization in Rhizopus oryzae cells immobilized within biomass support particles for use as whole-cell biocatalysts in biodiesel-fuel production. J. Biosci. Bioeng., 101(4): 328-333.
    CrossRef    PMid:16716941    
11. Hama, S., H. Yamaji, T. Fukumizu, T. Numata, S. Tamalampudi, A. Kondo, H. Noda and H. Fukuda, 2007. Biodiesel-fuel production in a packed-bed reactor using lipase-producing Rhizopus oryzae cells immobilized within biomass support particles. Biochem. Eng. J., 34(3): 273-278.
    CrossRef    
12. Hama, S., A. Yoshida, K. Nakashima, H. Noda, H. Fukuda and A. Kondo, 2010. Surfactant-modified yeast whole-cell biocatalyst displaying lipase on cell surface for enzymatic production of structured lipids in organic media. Appl. Microbiol. Biot., 87(2): 537-543.
    CrossRef    PMid:20336291    
13. Ikemura, T., 1985. Codon usage and tRNA content in unicellular and multicellular organisms. Mol. Biol. Evol., 2(1): 13-34.
    PMid:3916708    Direct Link
14. Kaieda, M., T. Samukawa, T. Matsumoto, K. Ban, A. Kondo, Y. Shimada, H. Noda, F. Nomoto, K. Ohtsuka, E. Izumoto and H. Fukuda, 1999. Biodiesel fuel production from plant oil catalyzed by Rhizopus oryzae lipase in a water-containing system without an organic solvent. J. Biosci. Bioeng., 88(6): 627-631.
    CrossRef    Direct Link
15. Kato, M., J. Fuchimoto, T. Tanino, A. Kondo, H. Fukuda and M. Ueda, 2007. Preparation of a whole-cell biocatalyst of mutated Candida antaretica Lipase B (mCALB) by a yeast molecular display system and its practical properties. Appl. Microbiol. Biot., 75(3): 549-555.
    CrossRef    PMid:17262207    
16. Kondo, A. and M. Ueda, 2004. Yeast cell-surface display-applications of molecular display. Appl. Microbiol. Biot., 64(1): 28-40.
    CrossRef    PMid:14716465    
17. Lee, D.H., J.M. Kim, H.Y. Shin, S.W. Kang and S.W. Kim, 2006. Biodiesel production using a mixture of immobilized Rhizopus oryzae and Candida rugosa lipases. Biotechnol. Bioproc. E., 11(6): 522-525.
    CrossRef    
18. Lithwick, G. and H. Margalit, 2003. Hierarchy of sequence-dependent features associated with prokaryotic translation. Genome Res., 13(12): 2665-2673.
    CrossRef    PMid:14656971 PMCid:PMC403808    
19. Matsumoto, T., S. Takahashi, M. Kaieda, M. Ueda, A. Tanaka, H. Fukuda and A. Kondo, 2001. Yeast whole-cell biocatalyst constructed by intracellular overproduction of Rhizopus oryzae lipase is applicable to biodiesel fuel production. Appl. Microbiol. Biot., 57(4): 515-520.
    CrossRef    PMid:11762598    
20. Mingarro, I., C. Abad and L. Braco, 1995. Interfacial activation-based molecular bioimprinting of lipolytic enzymes. P. Natl. Acad. Sci. USA, 92(8): 3308-3312.
    CrossRef    PMid:7724558 PMCid:PMC42155    
21. Oda, M., M. Kaieda, S. Hama, H. Yamaji, A. Kondo, E. Izumoto and H. Fukuda, 2005. Facilitatory effect of immobilized lipase-producing Rhizopus oryzae cells on acyl migration in biodiesel-fuel production. Biochem. Eng. J., 23(1): 45-51.
    CrossRef    
22. Prim, N., A. Blanco, J. Martinez, F.I.J. Pastor and P. Diaz, 2000. estA, a gene coding for a cell-bound esterase from Paenibacillus sp. BP-23, is a new member of the bacterial subclass of type B carboxylesterases. Res. Microbiol., 151(4): 303-312.
    CrossRef    Direct Link
23. Shiraga, S., M. Kawakami and M. Ueda, 2004. Construction of combinatorial library of starch-binding domain of Rhizopus oryzae glucoamylase and screening of clones with enhanced activity by yeast display method. J. Mol. Catal. B-Enzym., 28(4-6): 229-234.
    CrossRef    Direct Link
24. Tamalampudi, S., M.R. Talukder, S. Hama, T. Numata, A. Kondo and H. Fukuda, 2008. Enzymatic production of biodiesel from Jatropha oil: A comparative study of immobilized-whole cell and commercial lipases as a biocatalyst. Biochem. Eng. J., 39(1): 185-189.
    CrossRef    
25. Yan, J.Y., Y.J. Yan, J.K. Yang, L. Xu and Y. Liu, 2009. Combined strategy for preparation of a bioimprinted Geotrichum sp. lipase biocatalyst effective in non-aqueous media. Process Biochem., 44(10): 1128-1132.
    CrossRef    
26. Yilmaz, E., 2002. Improving the application of microbial lipase by bio-imprinting at substrate-interfaces. World J. Microb. Biot., 18(1): 37-40.
    CrossRef    
27. Zou, J.T., V. Katavic, E.M. Giblin, D.L. Barton, S.L. MacKenzie, W.A. Keller, X. Hu and D.C. Taylor, 1997. Modification of seed oil content and acyl composition in the brassicaceae by expression of a yeast sn-2 acyltransferase gene. Plant Cell., 9(6): 909-923.
    CrossRef    PMid:9212466 PMCid:PMC156967    

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