Springer Nature is making Coronavirus research free. View research | View latest news | Sign up for updates

Improvement of a Candida antarctica lipase B-displaying yeast whole-cell biocatalyst and its application to the polyester synthesis reaction

Abstract

A Candida antarctica lipase B (CALB)-displaying yeast whole-cell biocatalyst was constructed with the integration of the CALB cell-surface display expression cassette in the yeast genome and cell fusion by mating. Lipase hydrolytic activity of the yeast whole-cell biocatalyst subsequently increased, in both a- and α-type yeast cells, with the number of copies of the CALB cell-surface display expression cassette introduced, and reached 43.6 and 32.2 U/g-dry cell at 168 h cultivation, respectively. The lipase hydrolytic activity of whole cells in diploid yeast cells containing eight copies of the CALB cell-surface expression cassette reached 117 U/g-dry cell, and this value is approximately ninefold higher than that of the previously reported haploid CALB cell-surface displaying yeast using a multi-copy plasmid (Tanino et al. Appl. Microbial Biotechnol 75:1319–1325, 2007). This improved novel CALB-displaying yeast whole-cell biocatalyst could repeatedly catalyze the polyester, polybutylene adipate, synthesis reaction, using adipic acid and 1, 4-butandiol as the monomer molecules, four times in succession. This is the first report of the polymer synthesis using enzyme displaying yeast as the catalyst. The ratios of cyclic compounds in the polybutylene adipates synthesized with the CALB-displaying yeast whole-cells were lower than that in the polybutylene adipate synthesized with conventional metal catalysis. From these results, it appears that the use of CALB-displaying yeast cells could be useful for the polyester synthesis reaction, with reduced by-product production.

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

Fig. 1
Fig. 2
Fig. 3

References

  1. Albertsson AC, Srivastava RK (2008) Recent development in enzyme-catalyzed ring-opening polymerization. Adv Drug Deliv Rev 60:1077–1093

  2. Binns F, Harffey P, Roberts SM, Taylor A (1997) Studies of lipase-catalyzed polyesterification of an unactivated diacid/diol system. J Polym Sci, Part A: Polym Chem 36:2069–2080

  3. Gai SA, Wittrup KD (2007) Yeast surface display for protein engineering and characterization. Curr Opin Struct Biol 17:467–473

  4. Hilker I, Schaafsma AEJ, Peters RAH, Heise A, Nijenhuis AJ (2007) Insight into lipase-catalyzed formation of macrocyclic oligoesters. European Polymer Journal 44:1441–1450

  5. Kanai T, Atomi H, Umemura K, Ueno H, Teranishi Y, Ueda M, Tanaka A (1996) A novel heterologous gene expression system in Saccharomyces cerevisiae using the isocitrate lyase gene promoter from Candida tropicalis. Appl Microbiol Biotechnol 44:759–765

  6. Kondo A, Ueda M (2004) Yeast cell-surface display—applications of molecular display. Appl Microbiol Biotechnol 64:28–40

  7. Kondo A, Sugihara S, Kuwahara M, Toshima K, Matsumura S (2008) Lipaase-catalyzed ring-opening polymerization of molecularly pure cyclic oligomers for use in synthesis and chemical recycling of aliphatic polyesters. Macromol Biosci 11:533–539

  8. Kumer A, Kalra B, Dekhterman A, Gross RA (2000) Efficient ring-opening polymerization and copolymerization of e-caprolactone and w-pentadecalactone catalyzed by Candida antarctica lipase B. Macromolecules 33:6303–6309

  9. Mahapatro A, Kalra B, Kumar A, Gross RA (2003) Lipase-catalyzed polycondensations: effect of substrates and solvent on chain formation dispersity, and end-group structure. Biomacromoleculers 4:544–551

  10. Mahapatro A, Kumer A, Gross RA (2004) Mild, solvent-free omega-hydroxy acid polycondensations catalyzed by Candida antarctica lipase B. Biomacromoleculers 5:62–68

  11. Matsumoto T, Fukuda H, Ueda M, Tanaka A, Kondo A (2002) Construction of yeast strains with high cell surface lipase activity by using novel display systems based on the Flo1p flocculation functional domain. Appl Environ Microbiol 68:4517–4522

  12. Mesiano AJ, Beckman EJ, Russell AJ (2000) Biocatalytic synthesis of fluorinated polyesters. Biotechnol Prog 16:64–68

  13. Mezoul G, Lalot T, Brigodiot M, Marechal E (1996) Enzyme-catalyzed synthesis of poly(1,6-hexanediyl isophthalate) and poly(1,6-hexanediyl terephthalate). Polym Bull 36:541–548

  14. Tanino T, Ohno T, Aoki T, Fukuda H, Kondo A (2007) Development of yeast cells displaying Candida antarctica lipase B and their application to ester synthesis reaction. Appl Microbial Biotechnol 75:1319–1325

  15. Ueda M, Tanaka A (2000a) Genetic immobilization of proteins on the yeast cell surface. Biotechnol Adv 18:121–140

  16. Ueda M, Tanaka A (2000b) Cell surface engineering of yeast: construction of arming yeast with biocatalyst. L Biosci Bioeng 90:125–136

  17. Ugolini S, Bruschi CV (1996) The red/white color assay in the yeast Saccharomyces cerevisiae: epistatic growth advantage of white ade8–18, ade2 cells over red ade2 cells. Curr Genet 30:485–492

  18. Umemura K, Atomi H, Kanai T, Teranishi Y, Ueda M, Tanaka A (1995) A novel promoter, derived from the isocitrate lyase gene of Candida tropicalis, inducible with acetate in Saccharomyces cerevisiae. Appl Microbiol Biotechnol 43:489–492

Download references

Author information

Correspondence to Akihiko Kondo.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Tanino, T., Aoki, T., Chung, W. et al. Improvement of a Candida antarctica lipase B-displaying yeast whole-cell biocatalyst and its application to the polyester synthesis reaction. Appl Microbiol Biotechnol 82, 59–66 (2009). https://doi.org/10.1007/s00253-008-1764-z

Download citation

Keyword

  • Cell-surface display
  • Candida antarctica
  • lipase B
  • Polyester
  • Synthesis
  • Diploid