DOI QR코드

DOI QR Code

Metabolic Engineering of Escherichia coli for Production of Polyhydroxyalkanoates with Hydroxyvaleric Acid Derived from Levulinic Acid

  • Kim, Doyun (Department of Biomedical Engineering, Ulsan National Institute of Science and Technology (UNIST)) ;
  • Lee, Sung Kuk (Department of Biomedical Engineering, Ulsan National Institute of Science and Technology (UNIST))
  • 투고 : 2021.08.16
  • 심사 : 2021.10.12
  • 발행 : 2022.01.28

초록

Polyhydroxyalkanoates (PHAs) are emerging as alternatives to plastics by replacing fossil fuels with renewable raw substrates. Herein, we present the construction of engineered Escherichia coli strains to produce short-chain-length PHAs (scl-PHAs), including the monomers 4-hydroxyvalerate (4HV) and 3-hydroxyvalerate (3HV) produced from levulinic acid (LA). First, an E. coli strain expressing genes (lvaEDABC) from the LA metabolic pathway of Pseudomonas putida KT2440 was constructed to generate 4HV-CoA and 3HV-CoA. Second, both PhaAB enzymes from Cupriavidus necator H16 were expressed to supply 3-hydroxybutyrate (3HB)-CoA from acetyl-CoA. Finally, PHA synthase (PhaCCv) from Chromobacterium violaceum was introduced for the subsequent polymerization of these three monomers. The resulting E. coli strains produced four PHAs (w/w% of dry cell weight): 9.1 wt% P(4HV), 1.7 wt% P(3HV-co-4HV), 24.2 wt% P(3HB-co-4HV), and 35.6 wt% P(3HB-co-3HV-co-4HV).

키워드

과제정보

This work has supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MIST) (2020R1A4A1018332) and granted by innovative science project in 2020 of The circle foundation

참고문헌

  1. Reinecke F, Steinbuchel A. 2009. Ralstonia eutropha strain H16 as model organism for PHA metabolism and for biotechnological production of technically interesting biopolymers. Microb. Physiol. 16: 91-108. https://doi.org/10.1159/000142897
  2. Yu J. 2007. Microbial production of bioplastics from renewable resources, pp. 585-610. Bioprocessing for value-added products from renewable resources, Ed. Elsevier,
  3. Pettinari MJ, Vazquez GJ, Silberschmidt D, Rehm B, Steinbuchel A, Mendez BS. 2001. Poly (3-hydroxybutyrate) synthesis genes in Azotobacter sp. strain FA8. Appl. Environ. Microbiol. 67: 5331-5334. https://doi.org/10.1128/AEM.67.11.5331-5334.2001
  4. Lutke-Eversloh T, Steinbuchel A. 2004. Microbial polythioesters. Macromol. Biosci. 4: 165-174. https://doi.org/10.1002/mabi.200300084
  5. Napper IE, Thompson RC. 2019. Environmental deterioration of biodegradable, oxo-biodegradable, compostable, and conventional plastic carrier bags in the sea, soil, and open-air over a 3-year period. Environ. Sci. Technol. 53: 4775-4783. https://doi.org/10.1021/acs.est.8b06984
  6. Bhatia SK, Gurav R, Choi T-R, Jung H-R, Yang S-Y, Moon Y-M, et al. 2019. Bioconversion of plant biomass hydrolysate into bioplastic (polyhydroxyalkanoates) using Ralstonia eutropha 5119. Bioresour. Technol. 271: 306-315. https://doi.org/10.1016/j.biortech.2018.09.122
  7. Lemoigne M. 1926. Products of dehydration and of polymerization of β-hydroxybutyric acid. Bull. Chem. Soc. Japan 8: 770-782.
  8. Muzaiyanah AR, Amirul AA. 2013. Studies on the microbial synthesis and characterization of polyhydroxyalkanoates containing 4-hydroxyvalerate using γ-valerolactone. Appl. Biochem. Biotechnol. 170: 1194-1215. https://doi.org/10.1007/s12010-013-0247-6
  9. Raza ZA, Abid S, Banat IM. 2018. Polyhydroxyalkanoates: Characteristics, production, recent developments and applications. Int. Biodeterior. Biodegradation 126: 45-56. https://doi.org/10.1016/j.ibiod.2017.10.001
  10. Mozejko-Ciesielska J, Kiewisz R. 2016. Bacterial polyhydroxyalkanoates: Still fabulous? Microbiol. Res. 192: 271-282. https://doi.org/10.1016/j.micres.2016.07.010
  11. Sheu D-S, Chen Y-LL, Jhuang W-J, Chen H-Y, Jane W-N. 2018. Cultivation temperature modulated the monomer composition and polymer properties of polyhydroxyalkanoate synthesized by Cupriavidus sp. L7L from levulinate as sole carbon source. Int. J. Biol. Macromol. 118: 1558-1564. https://doi.org/10.1016/j.ijbiomac.2018.06.193
  12. Gahlawat G, Soni SK. 2017. Valorization of waste glycerol for the production of poly (3-hydroxybutyrate) and poly (3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer by Cupriavidus necator and extraction in a sustainable manner. Bioresour. Technol. 243: 492-501. https://doi.org/10.1016/j.biortech.2017.06.139
  13. Schmack G, Gorenflo V, Steinbuchel A. 1998. Biotechnological production and characterization of polyesters containing 4-hydroxyvaleric acid and medium-chain-length hydroxyalkanoic acids. Macromolecules 31: 644-649. https://doi.org/10.1021/ma970864d
  14. Lee W-H, Loo C-Y, Nomura CT, Sudesh K. 2008. Biosynthesis of polyhydroxyalkanoate copolymers from mixtures of plant oils and 3-hydroxyvalerate precursors. Bioresour. Technol. 99: 6844-6851. https://doi.org/10.1016/j.biortech.2008.01.051
  15. Valentin HE, Steinbuchel A. 1995. Accumulation of poly(3-hydroxybutyric acid-co-3-hydroxyvaleric acid-co-4-hydroxyvaleric acid) by mutants and recombinant strains of Alcaligenes eutrophus. J. Environ. Polymer Degradation 3: 169-175. https://doi.org/10.1007/BF02068468
  16. Koller M, Hesse P, Fasl H, Stelzer F, Braunegg G. 2017. Study on the effect of levulinic acid on whey-based biosynthesis of Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) by Hydrogenophaga pseudoflava. Appl. Food Biotechnol. 4: 65-78.
  17. Novackova I, Kucera D, Porizka J, Pernicova I, Sedlacek P, Koller M, et al. 2019. Adaptation of Cupriavidus necator to levulinic acid for enhanced production of P(3HB-co-3HV) copolyesters. Biochem. Eng. J. 151: 107350. https://doi.org/10.1016/j.bej.2019.107350
  18. Cha D, Ha HS, Lee SK. 2020. Metabolic engineering of Pseudomonas putida for the production of various types of short-chain-length polyhydroxyalkanoates from levulinic acid. Bioresour. Technol. 309: 123332. https://doi.org/10.1016/j.biortech.2020.123332
  19. Bozell JJ, Moens L, Elliott DC, Wang Y, Neuenscwander GG, Fitzpatrick SW, et al. 2000. Production of levulinic acid and use as a platform chemical for derived products. Resour. Conserv. Recycl. 28: 227-239. https://doi.org/10.1016/S0921-3449(99)00047-6
  20. Favaro L, Basaglia M, Casella S. 2019. Improving polyhydroxyalkanoate production from inexpensive carbon sources by genetic approaches: a review. Biofuels Bioprod. Biorefining 13: 208-227. https://doi.org/10.1002/bbb.1944
  21. Kidwell J, Valentin HE, Dennis D. 1995. Regulated expression of the Alcaligenes eutrophus pha biosynthesis genes in Escherichia coli. Appl. Environ. Microbiol. 61: 1391-1398. https://doi.org/10.1128/aem.61.4.1391-1398.1995
  22. Langenbach S, Rehm BHA, Steinbuchel A. 1997. Functional expression of the PHA synthase gene phaC1 from Pseudomonas aeruginosa in Escherichia coli results in poly(3-hydroxyalkanoate) synthesis. FEMS Microbiol. Lett. 150: 303-309. https://doi.org/10.1016/S0378-1097(97)00142-0
  23. Ren Q, Sierro N, Kellerhals M, Kessler B, Witholt B. 2000. Properties of engineered poly-3-hydroxyalkanoates produced in recombinant Escherichia coli strains. Appl. Environ. Microbiol. 66: 1311-1320. https://doi.org/10.1128/AEM.66.4.1311-1320.2000
  24. Ren Q, Beilen JBv, Sierro N, Zinn M, Kessler B, Witholt B. 2005. Expression of PHA polymerase genes of Pseudomonas putida in Escherichia coli and its effect on PHA formation. Antonie Van Leeuwenhoek 87: 91-100. https://doi.org/10.1007/s10482-004-1360-x
  25. Wang Q, Yu H, Xia Y, Kang Z, Qi Q. 2009. Complete PHB mobilization in Escherichia coli enhances the stress tolerance: a potential biotechnological application. Microb. Cell Fact. 8: 1-9. https://doi.org/10.1186/1475-2859-8-1
  26. Rand JM, Pisithkul T, Clark RL, Thiede JM, Mehrer CR, Agnew DE, et al. 2017. A metabolic pathway for catabolizing levulinic acid in bacteria. Nat. Microbiol. 2: 1624-1634. https://doi.org/10.1038/s41564-017-0028-z
  27. Lee TS, Krupa RA, Zhang F, Hajimorad M, Holtz WJ, Prasad N, et al. 2011. BglBrick vectors and datasheets: A synthetic biology platform for gene expression. J. Biol. Eng. 5: 12. https://doi.org/10.1186/1754-1611-5-12
  28. Gibson DG, Young L, Chuang R-Y, Venter JC, Hutchison CA, Smith HO. 2009. Enzymatic assembly of DNA molecules up to several hundred kilobases. Nat. Methods 6: 343-345. https://doi.org/10.1038/nmeth.1318
  29. Reis AC, Salis HM. 2020. An automated model test system for systematic development and improvement of gene expression models. ACS Synth. Biol. 9: 3145-3156. https://doi.org/10.1021/acssynbio.0c00394
  30. Gorenflo V, Schmack G, Vogel R, Steinbuchel A. 2001. Development of a process for the biotechnological large-scale production of 4-hydroxyvalerate-containing polyesters and characterization of their physical and mechanical properties. Biomacromolecules 2: 45-57. https://doi.org/10.1021/bm0000992
  31. Juengert JR, Bresan S, Jendrossek D. 2018. Determination of polyhydroxybutyrate (PHB) content in Ralstonia eutropha using gas chromatography and nile red staining. Bio Protoc. 8: e2748.
  32. Pena C, Castillo T, Garcia A, Millan M, Segura D. 2014. Biotechnological strategies to improve production of microbial poly-(3-hydroxybutyrate): a review of recent research work. Microb. Biotechnol. 7: 278-293. https://doi.org/10.1111/1751-7915.12129
  33. Matsumoto Ki, Yamada M, Leong CR, Jo S-J, Kuzuyama T, Taguchi S. 2011. A new pathway for poly (3-hydroxybutyrate) production in Escherichia coli and Corynebacterium glutamicum by functional expression of a new acetoacetyl-coenzyme A synthase. Biosci. Biotechnol. Biochem. 75: 364-366. https://doi.org/10.1271/bbb.100682
  34. Snell KD, Feng F, Zhong L, Martin D, Madison LL. 2002. YfcX enables medium-chain-length poly (3-hydroxyalkanoate) formation from fatty acids in recombinant Escherichia coli fadB strains. J. Bacteriol. 184: 5696-5705. https://doi.org/10.1128/JB.184.20.5696-5705.2002
  35. Liu S-J, Steinbuchel A. 2000. A novel genetically engineered pathway for synthesis of poly(hydroxyalkanoic acids) in Escherichia coli. Appl. Environ. Microbiol. 66: 739-743. https://doi.org/10.1128/AEM.66.2.739-743.2000
  36. Kim BS, Lee SY. 2000. Production of poly (3-hydroxybutyrate) from inexpensive substrates. Enzyme Microb. Technol. 27: 774-777. https://doi.org/10.1016/S0141-0229(00)00299-4
  37. Kim D, Sathesh-Prabu C, JooYeon Y, Lee SK. 2019. High-level production of 4-hydroxyvalerate from levulinic acid via whole-cell biotransformation decoupled from cell metabolism. J. Agric. Food Chem. 67: 10678-10684. https://doi.org/10.1021/acs.jafc.9b04304
  38. Steinbuchel A, Debzi E-M, Marchessault RH, Timm A. 1993. Synthesis and production of poly (3-hydroxyvaleric acid) homopolyester by Chromobacterium violaceum. Appl. Microbiol. Biotechnol. 39: 443-449. https://doi.org/10.1007/BF00205030
  39. Tripathi L, Wu L-P, Dechuan M, Chen J, Wu Q, Chen G-Q. 2013. Pseudomonas putida KT2442 as a platform for the biosynthesis of polyhydroxyalkanoates with adjustable monomer contents and compositions. Bioresour. Technol. 142: 225-231. https://doi.org/10.1016/j.biortech.2013.05.027
  40. Aldor IS, Keasling JD. 2003. Process design for microbial plastic factories: metabolic engineering of polyhydroxyalkanoates. Curr. Opin. Biotechnol. 14: 475-483. https://doi.org/10.1016/j.copbio.2003.09.002
  41. Blattner FR, Plunkett G, Bloch CA, Perna NT, Burland V, Riley M, et al. 1997. The complete genome sequence of Escherichia coli K-12. Science 277: 1453-1462. https://doi.org/10.1126/science.277.5331.1453
  42. Durfee T, Nelson R, Baldwin S, Plunkett G, Burland V, Mau B, et al. 2008. The complete genome sequence of Escherichia coli DH10b: insights into the biology of a laboratory workhorse. J. Bacteriol. 190: 2597-2606. https://doi.org/10.1128/JB.01695-07