Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
권호 표지
DOI QR코드

DOI QR Code

Synthesis of Isoamyl Fatty Acid Ester, a Flavor Compound, by Immobilized Rhodococcus Cutinase

  • Ye Won Jeon (Division of Biotechnology, The Catholic University of Korea) ;
  • Ha Min Song (Division of Biotechnology, The Catholic University of Korea) ;
  • Ka Yeong Lee (Division of Biotechnology, The Catholic University of Korea) ;
  • Yeong A Kim (Division of Biotechnology, The Catholic University of Korea) ;
  • Hyung Kwoun Kim (Division of Biotechnology, The Catholic University of Korea)
  • Received : 2024.02.21
  • Accepted : 2024.04.05
  • Published : 2024.06.28
Download PDF
⟨ Previous Next ⟩

Abstract

Isoamyl fatty acid esters (IAFEs) are widely used as fruity flavor compounds in the food industry. In this study, various IAFEs were synthesized from isoamyl alcohol and various fatty acids using a cutinase enzyme (Rcut) derived from Rhodococcus bacteria. Rcut was immobilized on methacrylate divinylbenzene beads and used to synthesize isoamyl acetate, butyrate, hexanoate, octanoate, and decanoate. Among them, Rcut synthesized isoamyl butyrate (IAB) most efficiently. Docking model studies showed that butyric acid was the most suitable substrate in terms of binding energy and distance from the active site serine (Ser114) γ-oxygen. Up to 250 mM of IAB was synthesized by adjusting reaction conditions such as substrate concentration, reaction temperature, and reaction time. When the enzyme reaction was performed by reusing the immobilized enzyme, the enzyme activity was maintained at least six times. These results demonstrate that the immobilized Rcut enzyme can be used in the food industry to synthesize a variety of fruity flavor compounds, including IAB.

Keywords

Acknowledgement

This research was supported by Korea Polar Research Institute (PE23150) and funded by the Ministry of Oceans and Fisheries.

References

  1. Chen S, Tong X, Woodard RW, Du G, Wu J, Chen J. 2008. Identification and characterization of bacterial cutinase. J. Biol. Chem. 283: 25854-25862. https://doi.org/10.1074/jbc.M800848200
  2. Martinez A, Maicas S. 2021. Cutinases: Characteristics and insights in industrial production. Catalysts 11: 1194.
  3. Vazquez-Alcantara L, Oliart-Ros RM, Garcia-Borquez A, Pena-Montes C. 2021. Expression of a cutinase of Moniliophthora roreri with polyester and PET-plastic residues degradation activity. Microbiol. Spectr. 9: e0097621.
  4. Yan ZF, Wang L, Xia W, Liu ZZ, Gu LT, Wu J. 2021. Synergistic biodegradation of poly (ethylene terephthalate) using Microbacterium oleivorans and Thermobifida fusca cutinase. Appl. Microbiol. Biotechnol. 105: 4551-4560. https://doi.org/10.1007/s00253-020-11067-z
  5. Won SJ, Yim JH, Kim HK. 2023. Synthesis of short-chain alkyl butyrate through esterification reaction using immobilized Rhodococcus cutinase and analysis of substrate specificity through molecular docking. J. Microbiol. Biotechnol. 33: 268-276. https://doi.org/10.4014/jmb.2211.11022
  6. Lai J, Huang H, Lin M, Xu Y, Li X, Sun B. 2023. Enzyme catalyzes ester bond synthesis and hydrolysis: the key step for sustainable usage of plastics. Front. Microbiol. 13: 1113705.
  7. Purdy RE, Kolattukudy PE. 1975. Hydrolysis of plant cuticle by plant pathogens. purification, amino acid composition, and molecular weight of two isozymes of cutinase and a nonspecific esterase from Fusarium solani f. pisi. Biochemistry 14: 2824-2831. https://doi.org/10.1021/bi00684a006
  8. Xiuhong L, Huibin Z. 2023. Biotechnological application of cutinase: a powerful tool in synthetic biology. SynBio. 1: 54-64. https://doi.org/10.3390/synbio1010004
  9. de Barros DPC, Lemos F, Fonseca LP, Cabral JMS. 2010. Kinetic cutinase-catalyzed esterification of caproic acid in organic solvent system. J. Mol. Catal. B Enzym. 66: 285-293. https://doi.org/10.1016/j.molcatb.2010.06.005
  10. Martinez C, De Geus P, Lauwereys M, Matthyssens G, Cambillau C. 1992. Fusarium solani cutinase is a lipolytic enzyme with a catalytic serine accessible to solvent. Nature 356: 615-618. https://doi.org/10.1038/356615a0
  11. Liu Z, Gosser Y, Baker PJ, Ravee Y, Lu Z, Alemu G, et al. 2009. Structural and functional studies of Aspergillus oryzae cutinase: enhanced thermostability and hydrolytic activity of synthetic ester and polyester degradation. J. Am. Chem. Soc. 131: 15711-15716. https://doi.org/10.1021/ja9046697
  12. Dimarogona M, Nikolaivits E, Kanelli M, Christakopoulos P, Sandgren M, Topakas E. 2015. Structural and functional studies of a Fusarium oxysporum cutinase with polyethylene terephthalate modification potential. Biochim. Biophys. Acta 1850: 2308-2317. https://doi.org/10.1016/j.bbagen.2015.08.009
  13. Kold D, Dauter Z, Laustsen AK, Brzozowski AM, Turkenburg JP, Nielsen AD, et al. 2014. Thermodynamic and structural investigation of the specific SDS binding of Humicola insolens cutinase. Protein Sci. 23: 1023-1035. https://doi.org/10.1002/pro.2489
  14. Roussel A, Amara S, Nyyssola A, Mateos-Diaz E, Blangy S, Kontkanen H, et al. 2014. A Cutinase from Trichoderma reesei with a lid-covered active site and kinetic properties of true lipases. J. Mol. Biol. 426: 3757-3772. https://doi.org/10.1016/j.jmb.2014.09.003
  15. Duan X, Liu Y, You X, Jiang Z, Yang S, Yang S. 2017. High-level expression and characterization of a novel cutinase from Malbranchea cinnamomea suitable for butyl butyrate production. Biotechnol. Biofuels 10: 223.
  16. Kitadokoro K, Thumarat U, Nakamura R, Nishimura K, Karatani H, Suzuki H, et al. 2012. Crystal structure of cutinase Est119 from Thermobifida alba AHK119 that can degrade modified polyethylene terephthalate at 1.76A resolution. Polym. Degrad. Stab. 97: 771-775. https://doi.org/10.1016/j.polymdegradstab.2012.02.003
  17. Roth C, Wei R, Oeser T, Then J, Follner C, Zimmermann W, et al. 2014. Structural and functional studies on a thermostable polyethylene terephthalate degrading hydrolase from Thermobifida fusca. Appl. Microbiol. Biotechnol. 98: 7815-7823. https://doi.org/10.1007/s00253-014-5672-0
  18. Su A, Kiokekli S, Naviwala M, Shirke AN, Pavlidis IV, Gross RA. 2020. Cutinases as stereoselective catalysts: specific activity and enantioselectivity of cutinases and lipases for menthol and its analogs. Enzyme Microb. Technol. 133.
  19. Won SJ, Yim JH, Kim HK. 2022. Functional production, characterization, and immobilization of a cold-adapted cutinase from Antarctic Rhodococcus sp. Protein Expr. Purif. 195-196: 106077
  20. Xu Y, Wang X, Liu X, Li X, Zhang C, Li W, et al. 2021. Discovery and development of a novel short-chain fatty acid ester synthetic biocatalyst under aqueous phase from Monascus purpureus isolated from Baijiu. Food Chem. 338: 128025.
  21. Vandamme EJ. Soetaert W. 2002. Bioflavours and fragrances via fermentation and biocatalysis. J. Chem. Technol. Biotechnol. 77: 1323-1332. https://doi.org/10.1002/jctb.722
  22. Khan NR, Rathod VK. 2015. Enzyme catalyzed synthesis of cosmetic esters and its intensification: a review. Process Biochem. 50: 1793-1806. https://doi.org/10.1016/j.procbio.2015.07.014
  23. Muley AB, Chaudhari SA, Bankar SB, Singhal RS. 2019. Stabilization of cutinase by covalent attachment on magnetic nanoparticles and improvement of its catalytic activity by ultrasonication. Ultrason. Sonochem. 55: 174-185. https://doi.org/10.1016/j.ultsonch.2019.02.019
  24. de Barros DP, Azevedo AM, Cabral JM, Fonseca LP. 2012. Optimization of flavor esters synthesis by Fusarium solani pisi cutinase. J. Food Biochem. 36: 275-284. https://doi.org/10.1111/j.1745-4514.2010.00535.x
  25. Dutta K, Dasu VV. 2011. Synthesis of short chain alkyl esters using cutinase from Burkholderia cepacia NRRL B2320. J. Mol. Catal. B Enzym. 72: 150-156. https://doi.org/10.1016/j.molcatb.2011.05.013
  26. Su L, Hong R, Guo X, Wu J, Xia Y. 2016. Short-chain aliphatic ester synthesis using Thermobifida fusca cutinase. Food Chem. 206: 131-136. https://doi.org/10.1016/j.foodchem.2016.03.051
  27. Thomas S. 2002. The hydrogen bond in the solid state. Angew. Chem. Int. Ed. 41: 48-76. https://doi.org/10.1002/1521-3773(20020104)41:1<48::AID-ANIE48>3.0.CO;2-U
  28. Nikolaivits E, Makris G, Topakas E. 2017. Immobilization of a cutinase from Fusarium oxysporum and application in pineapple flavor synthesis. J. Agric. Food Chem. 65: 3505-3511. https://doi.org/10.1021/acs.jafc.7b00659
  29. Lai DT, O'Connor CJ. 1999. Studies on synthesis of short chain alkyl esters catalyzed by goat pregastric lipase. J. Mol. Catal. B Enzym. 6: 411-420. https://doi.org/10.1016/S1381-1177(99)00007-7
  30. Romero MD, Calvo L, Alba C, Daneshfar A. 2007. A kinetic study of isoamyl acetate synthesis by immobilized lipase-catalyzed acetylation in n-hexane. J. Biotechnol. 127: 269-277. https://doi.org/10.1016/j.jbiotec.2006.07.009
  31. Chaudhari SA, Singhal RS. 2017. A strategic approach for direct recovery and stabilization of Fusarium sp. ICT SAC1 cutinase from solid state fermented broth by carrier free cross-linked enzyme aggregates. nt. J. Biol. Macromol. 98: 610-621. https://doi.org/10.1016/j.ijbiomac.2017.02.033
  32. Ali SH, Tarakmah A, Merchant SQ, Al-Sahhaf T. 2007. Synthesis of esters: development of the rate expression for the Dowex 50 Wx8-400 catalyzed esterification of propionic acid with 1-propanol. Chem. Eng. Sci. 62: 3197-3217. https://doi.org/10.1016/j.ces.2007.03.017
  33. Romero MD, Calvo L, Alba C, Daneshfar A. 2007. A kinetic study of isoamyl acetate synthesis by immobilized lipase-catalyzed acetylation in n-hexane. J. Biotechnol. 127: 269-277. https://doi.org/10.1016/j.jbiotec.2006.07.009
  34. Paludo N, Alves JS, Altmann C, Ayub MA, Fernandez-Lafuente R, Rodrigues RC. 2015. The combined use of ultrasound and molecular sieves improves the synthesis of ethyl butyrate catalyzed by immobilized Thermomyces lanuginosus lipase. Ultrason. Sonochem. 22: 89-94. https://doi.org/10.1016/j.ultsonch.2014.05.004