Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Mcl-PHAs Produced by Pseudomonas sp. Gl01 Using Fed-Batch Cultivation with Waste Rapeseed Oil as Carbon Source

  • Mozejko, Justyna (Department of Environmental Biotechnology, University of Warmia and Mazury in Olsztyn) ;
  • Wilke, Andreas (Department of Mechanical and Process Engineering, University of Applied Sciences Offenburg) ;
  • Przybylek, Grzegorz (Department of Environmental Biotechnology, University of Warmia and Mazury in Olsztyn) ;
  • Ciesielski, Slawomir (Department of Environmental Biotechnology, University of Warmia and Mazury in Olsztyn)
  • Received : 2011.06.20
  • Accepted : 2011.11.05
  • Published : 2012.03.28
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Abstract

The present study describes medium-chain-length polyhydroxyalkanoates (mcl-PHAs) production by the Pseudomonas Gl01 strain isolated from mixed microbial communities utilized for PHAs synthesis. A two-step fed-batch fermentation was conducted with glucose and waste rapeseed oil as the main carbon source for obtaining cell growth and mcl-PHAs accumulation, respectively. The results show that the Pseudomonas Gl01 strain is capable of growing and accumulating mcl-PHAs using a waste oily carbon source. The biomass value reached 3.0 g/l of CDW with 20% of PHAs content within 48 h of cultivation. The polymer was purified from lyophilized cells and analyzed by gas chromatography (GC). The results revealed that the monomeric composition of the obtained polyesters depended on the available substrate. When glucose was used in the growth phase, 3-hydroxyundecanoate and 3-hydroxydodecanoate were found in the polymer composition, whereas in the PHAs-accumulating stage, the Pseudomonas Gl01 strain synthesized mcl-PHAs consisting mainly of 3-hydroxyoctanoate and 3-hydroxydecanoate. The transcriptional analysis using reverse-transcription real-time PCR reaction revealed that the phaC1 gene could be transcribed simultaneously to the phaZ gene.

Keywords

References

  1. Anderson, A. J. and E. A. Dawes. 1990. Occurence, metabolism, metabolic role, and industrial uses of bacterial polyhydroxyalkanoates. Microbiol. Rev. 54: 450-472.
  2. American Public Health Association. 1992. Standard Methods for the Examination of Water and Wastewater, 20th Ed. American Public Health Association, Washington.
  3. Braunegg, G., B. Sonnleitner, and R. M. Lafferty. 1978. A rapid gas chromatographic method for the determination of poly-bhydroxybutyric acid in microbial biomass. Eur. J. Appl. Microbiol. Biotechnol. 6: 29-37. https://doi.org/10.1007/BF00500854
  4. Carnicero, D., M. Fernández-Valverde, L. M. Canedo, C. Schleissner, and J. M. Luengo. 1997. Octanoic acid uptake in Pseudomonas putida U. FEMS Microbiol. Lett. 149: 51-58. https://doi.org/10.1111/j.1574-6968.1997.tb10307.x
  5. Ciesielski, S., J. Mo ejko, and G. Przyby ek. 2010. The influence of nitrogen limitation on mcl-PHA synthesis by two newly isolated strains of Pseudomonas sp. J. Ind. Microbiol. Biotechnol. 37: 511-520. https://doi.org/10.1007/s10295-010-0698-5
  6. Ciesielski, S., T. Pokoj, and E. Klimiuk. 2010. Cultivationdependent and -independent characterization of microbial community producing polyhydroxyalkanoates from raw-glycerol. J. Microbiol. Biotechnol. 20: 853-861. https://doi.org/10.4014/jmb.0909.09038
  7. Diniz, S. C., M. K. Taciro, J. G. C. Gomez, and J. G. da Cruz Pradella. 2004. High-cell-density cultivation of Pseudomonas putida IPT 046 and medium-chain-length polyhydroxyalkanoates production from sugarcane carbohydrates. Appl. Biochem. Biotechnol. 119: 51-69. https://doi.org/10.1385/ABAB:119:1:51
  8. Durner, R., M. Zinn, B. Witholt, and T. Egli. 2001. Accumulation of poly[(R)-3-hydroxyalkanoates] in Pseudomonas oleovorans during growth in batch and chemostat culture with different carbon sources. Biotechnol. Bioeng. 72: 278-288. https://doi.org/10.1002/1097-0290(20010205)72:3<278::AID-BIT4>3.0.CO;2-G
  9. Furrer, P., R. Hany, D. Rentsch, A. Grubelnik, K. Ruth, S. Panke, and M. Zinn. 2007. Quantitative analysis of bacterial medium-chain-length poly([R]-3-hydroxyalkanoates) by gas chromatography. J. Chromatogr. A 1143: 199-206. https://doi.org/10.1016/j.chroma.2007.01.002
  10. Haba, E., J. Vidal-Mas, M. Bassas, M. J. Espuny, J. Llorens, and A. Manresa. 2007. Poly 3-(hydroxyalkanoates) produced from oily substrates by Pseudomonas aeruginosa 47T2 (NCBIM 40044): Effect of nutrients and incubation temperature on polymer composition. Biochem. Eng. J. 35: 99-106. https://doi.org/10.1016/j.bej.2006.11.021
  11. Hartmann, R., R. Hany, E. Pletscher, A. Ritter, B. Witholt, and M. Zinn. 2006. Tailor-made olefinic medium-chain-length poly[(R)-3-hydroxyalkanoates] by Pseudomonas putida GPo1: Batch versus chemostat production. Biotechnol. Bioeng. 93: 737-746. https://doi.org/10.1002/bit.20756
  12. Hoffman, N. and B. H. A. Rehm. 2004. Regulation of polyhydroxyalkanoate biosynthesis in Pseudomonas putida and Pseudomonas aeruginosa. FEMS Microbiol. Lett. 237: 1-7. https://doi.org/10.1111/j.1574-6968.2004.tb09671.x
  13. Huijberts, G. N. M. and G. Eggink. 1996. Production of poly(3-hydroxyalkanoates) by Pseudomonas putida KT2442 in continuous cultures. Appl. Microbiol. Biotechnol. 46: 233-239. https://doi.org/10.1007/s002530050810
  14. Huisman, G. W., E. Wonink, G. Koning, H. Preusting, and B. Witholt. 1992. Synthesis of poly(3-hydroxyalkanoates) by mutant and recombinant Pseudomonas strains. Appl. Microbiol. Biotechnol. 38: 1-5.
  15. Kim, G. J., I. Y. Lee, S. C. Yoon, Y. C. Shin, and Y. H. Park. 1997. Enhanced yield and a high production of medium-chainlength poly(3-hydroxyalkanoates) in a two-step fed-batch cultivation of Pseudomonas putida by combined use of glucose and octanoate. Enzyme Microb. Technol. 20: 500-505. https://doi.org/10.1016/S0141-0229(96)00179-2
  16. Lageveen, R. G., G. W. Huisman, H. Preusting, P. Ketelaar, G. Eggink, and B. Witholt. 1988. Formation of polyesters by Pseudomonas oleovorans: Effect of substrates on formation and composition of poly-(R)-3-hydroxyalkanoates and poly-(R)-3-hydroxyalkenoates. Appl. Environ. Microbiol. 54: 2924-2932.
  17. Lee, J., S. Y. Lee, S. Park, and A. P. Middelberg. 1999. Control of fed-batch fermentations. Biotechnol. Adv. 17: 29-48.
  18. Lee, S. Y. 1996. Bacterial polyhydroxyalkanoates. Biotechnol. Bioeng. 49: 1-14. https://doi.org/10.1002/(SICI)1097-0290(19960105)49:1<1::AID-BIT1>3.3.CO;2-1
  19. Lee, S. Y., H. H. Wong, J. Choi, S. H. Lee, S. C. Lee, and C. S. Han. 2000. Production of medium-chain-length polyhydroxyalkanoates by high-cell-density cultivation of Pseudomonas putida under phosphorus limitation. Biotechnol. Bioeng. 68: 466-470. https://doi.org/10.1002/(SICI)1097-0290(20000520)68:4<466::AID-BIT12>3.0.CO;2-T
  20. Livak, K. J. and T. D. Schmittgen. 2001. Analysis of relative gene expression data using real-time quantitative PCR and the $2^{-{\Delta}{\Delta}CT}$ method. Methods 25: 402-408. https://doi.org/10.1006/meth.2001.1262
  21. Lopez-Cuellar, M. R., J. Alba-Flores, J. N. Gracida-Rodriguez, and F. Perez-Guevara. 2011. Production of polyhydroxyalkanoates (PHAs) with canola oil as carbon source. Int. J. Biol. Macromol. 48: 74-80. https://doi.org/10.1016/j.ijbiomac.2010.09.016
  22. Ramsay, B. A., I. Saracovan, J. A. Ramsay, and R. H. Marchessault. 1991. Continuous production of long-side-chain poly-${\beta}$-hydroxyalkanoates by Pseudomonas oleovorans. Appl. Environ. Microbiol. 57: 625-629.
  23. Ramsay, B. A., I. Saracovan, J. A. Ramsay, and R. Marchessault. 1992. Effect of nitrogen limitation on long-side-chain poly-betahydroxyalkanoate synthesis by Pseudomonas resinovorans. Appl. Environ. Microbiol. 58: 744-746.
  24. Ren, Q., G. de Roo, B. Witholt, M. Zinn, and L. Thony-Meyer. 2010. Influence of growth stage on activities of polyhydroxyalkanoate (PHA) polymerase and PHA depolymerase in Pseudomonas putida U. BMC Microbiol. 10: 254-262. https://doi.org/10.1186/1471-2180-10-254
  25. Riesenberg, D. and R. Guthke. 1999. High-cell-density cultivation of microorganisms. Appl. Microbiol. Biotechnol. 51: 422-430. https://doi.org/10.1007/s002530051412
  26. Schlegel, H. G., G. Gottschalk, and R. von Bartha. 1961. Formation and utilization of poly-${\beta}$-hydroxybutyric acid by knallgas bacteria (Hydrogenomonas). Nature 191: 463-465. https://doi.org/10.1038/191463a0
  27. Silva-Queiroza, S. R., L. F. Silva, J. G. C. Pradella, E. M. Pereira, and J. G. C. Gomez. 2009. PHAMCL biosynthesis systems in Pseudomonas aeruginosa and Pseudomonas putida strains show differences on monomer specificities. J. Biotechnol. 143: 111-118. https://doi.org/10.1016/j.jbiotec.2009.06.014
  28. Solaiman, D. K., R. D. Ashby, and T. A. Foglia. 2000. Rapid and specific identification of medium-chain-length polyhydroxyalkanoate synthase gene by polymerase chain reaction. Appl. Microbiol. Biotechnol. 53: 690-694. https://doi.org/10.1007/s002530000332
  29. Sun, Z., J. A. Ramsay, M. Guay, and B. A. Ramsay. 2007. Automated feeding strategies for high-cell-density fed-batch cultivation of Pseudomonas putida KT2440. Appl. Microbiol. Biotechnol. 71: 423-431.
  30. Sun, Z., J. A. Ramsay, M. Guay, and B. A. Ramsay. 2007. Carbon-limited fed-batch production of medium-chain-length polyhydroxyalkanoates from nonanoic acid by Pseudomonas putida KT2440. Appl. Microbiol. Biotechnol. 74: 69-77. https://doi.org/10.1007/s00253-006-0655-4
  31. Sun, Z., J. A. Ramsay, M. Guay, and B. A. Ramsay. 2007. Increasing the yield of mcl-PHA from nonanoic acid by cofeeding glucose during the PHA accumulation stage in twostage fed-batch fermentations of Pseudomonas putida KT2440. J. Biotechnol. 132: 280-282. https://doi.org/10.1016/j.jbiotec.2007.02.023
  32. Sun, Z., J. A. Ramsay, M. Guay, and B. A. Ramsay. 2009. Enhanced yield of medium-chain-length polyhydroxyalkanoates from nonanoic acid by co-feeding glucose in carbon-limited, fed-batch culture. J. Biotechnol. 143: 262-267. https://doi.org/10.1016/j.jbiotec.2009.07.014

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