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Evaluating haloarchaeal culture media for ultrahigh-molecular-weight polyhydroxyalkanoate biosynthesis by Haloferax mediterranei

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Abstract

A series of culture media for haloarchaea were evaluated to optimize the production of ultrahigh-molecular-weight (UHMW) poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) by Haloferax mediterranei. Cells of H. mediterranei grew (> 1 g/L of dry cell weight) and accumulated PHBV upon flask cultivation in 10 medium types with neutral pH and NaCl concentration > 100 g/L. Molecular weight and compositional analysis revealed that the number-average molecular weight (Mn) of PHBV produced with six selected types of media ranged from 0.8 to 3.5 × 106 g/mol and the 3-hydroxyvalerate (3HV) composition ranged from 8 to 36 mol%. Cultivation in two NBRC media, 1214 and 1380, resulted in the production of PHBV with an Mn of more than 3.0 × 106 g/mol and a weight-average molecular weight of more than 5.0 × 106 g/mol, indicating the production of UHMW-PHBV. These culture media contained small amount of complex nutrients like yeast extract and casamino acids, suggesting that H. mediterranei likely produced UHMW-PHBV on poor nutrient condition. Haloferax mediterranei grown in NBRC medium 1380 produced PHBV with the highest 3HV composition. A solvent-cast film of UHMW-PHBV with 26.4 mol% 3HV produced from 1-L flask cultivation with NBRC medium 1380 was found to be flexible and semi-transparent. Thermal analysis of the UHMW-PHBV cast film revealed melting and glass-transition temperatures of 90.5 °C and − 2.7 °C, respectively.

Key points

Haloarchaeal culture media were evaluated to produce UHMW-PHBV by H. mediterranei.

UHMW-PHBV with varied molecular weight was produced dependent on culture media.

Semi-transparent film could be made from UHMW-PHBV with 26.4 mol% 3HV.

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References

  • Alsafadi D, Al-Mashaqbeh O, Mansour A, Alsaad M (2020) Optimization of nitrogen source supply for enhanced biosynthesis and quality of poly(3-hydroxybutyurate-co-3-hydroxyvalerate) by extremely halophilic archaeon Haloferax mediterranei. MicrobiologyOpen 9:e1055

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Anjum A, Zuber M, Zia KM, Noreen A, Anjum MN, Tabasum S (2016) Microbial production of polyhydroxyalkanoates (PHAs) and its copolymers: a review of recent advancements. Int J Biol Macromol 89:161–174

    Article  CAS  PubMed  Google Scholar 

  • Aoyagi Y, Yamashita K, Doi Y (2002) Thermal degradation of poly[(R)-3-hydroxybutyrate], poly[ε-caprolactone], and poly[(S)-lactide]. Polym Degrad Stab 76:53–59

    Article  CAS  Google Scholar 

  • Aoyagi Y, Doi Y, Iwata T (2003) Mechanical properties and highly ordered structure of ultra-high-molecular-weight poly[(R)-3-hydroxybutyrate] films: effects of annealing and two-step drawing. Polym Degrad Stab 79:209–216

    Article  CAS  Google Scholar 

  • Avella M, La Rota G, Martuscelli E, Raimo M, Sadocco P, Elegir G, Riva R (2000) Poly(3-hydroxybutyrate- co-3-hydroxyvalerate) and wheat straw fibre composites: thermal, mechanical properties and biodegradation behaviour. J Mater Sci 35:829–836

  • Bowers KJ, Wiegel J (2011) Temperature and pH optima of extremely halophilic archaea: a mini-review. Extremophiles 15:119–128

    Article  CAS  PubMed  Google Scholar 

  • Chen G-Q, Jiang X-R (2018) Next generation industrial biotechnology based on extremophilic bacteria. Curr Opin Biotechnol 50:94–100

    Article  CAS  PubMed  Google Scholar 

  • Don TM, Chen CW, Chan TH (2006) Preparation and characterization of poly(hydroxyalkanoate) from the fermentation of Haloferax mediterranei. J Biomater Sci Polym Ed 17:1425–1438

    Article  CAS  PubMed  Google Scholar 

  • Fernandez-Castillo R, Rodriguez-Valera F, Gonzalez-Ramos J, Ruiz-Berraquero F (1986) Accumulation of poly(β-hydroxybutyrate) by halobacteria. Appl Environ Microbiol 51:214–216

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ferre-Guell A, Winterburn J (2017) Production of the copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate) with varied composition using different nitrogen sources with Haloferax mediterranei. Extremophiles 21:1037–1047

    Article  CAS  PubMed  Google Scholar 

  • Ferre-Guell A, Winterburn J (2018) Biosynthesis and characterization of polyhydroxyalkanoates with controlled composition and microstructure. Biomacromolecules 19:996–1005

    Article  CAS  Google Scholar 

  • Han J, Hou J, Zhang F, Ai G, Li M, Cai S, Liu H, Wang L, Wang Z, Zhang S, Cai L (2013) Multiple propionyl coenzyme A-supplying pathways for production of the bioplastic poly(3-hydroxybutyrate-co-3-hydroxyvalerate) in Haloferax mediterranei. Appl Environ Microbiol 79:2922–2931

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Han J, Wu LP, Hou J, Zhao D, Xiang H (2015) Biosynthesis, characterization, and hemostasis potential of tailor-made poly(3-hydroxybutyrate-co-3-hydroxyvalerate) produced by Haloferax mediterranei. Biomacromolecules 16:578–588

    Article  CAS  Google Scholar 

  • Hiroe A, Tsuge K, Nomura CT, Itaya M, Tsuge T (2012) Rearrangement of gene order in the phaCAB operon leads to effective production of ultrahigh-molecular-weight poly[(R)-3-hydroxybutyrate] in genetically engineered Escherichia coli. Appl Environ Microbiol 78:3177–3184

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ino K, Sato S, Ushimaru K, Saika A, Fukuoka T, Ohshiman K, Morita T (2020) Mechanical properties of cold-drawn films of ultrahigh-molecular-weight poly(3-hydroxybutyrate-co-3-hydroxyvalerate) produced by Haloferax mediterranei. Polym J 52:1299–1306

    Article  CAS  Google Scholar 

  • Iwata T (2005) Strong fibers and films of microbial polyesters. Macromol Biosci 5:689–701

    Article  CAS  PubMed  Google Scholar 

  • Iwata T, Doi Y (2005) Mechanical properties of uniaxially cold-drawn films of poly[(R)-3-hydroxybutyrate] and its copolymers. Macromol Symp 224:11–19

    Article  CAS  Google Scholar 

  • Iwata T, Tsunoda K, Aoyagi Y, Kusaka S, Yonezawa N, Doi Y (2003) Mechanical properties of uniaxially cold-drawn films of poly([R]-3-hydroxybutyrate). Polym Degrad Stab 79:217–224

    Article  CAS  Google Scholar 

  • Jendroseek D, Pfeiffer D (2014) New insights in the formation of polyhydroxyalkanoate granules (carbonosomes) and novel functions of poly(3-hydroxybutyrate). Environ Microbiol 16:2357–2373

    Article  CAS  Google Scholar 

  • Kamiya N, Yamamoto Y, Inoue Y, Chujo R, Doi Y (1989) Microstructure of bacterially synthesized poly(3-hydroxybutyrate-co-3-hydroxyvalerate). Macromolecules 22:1676–1682

    Article  CAS  Google Scholar 

  • Kato M, Bao HJ, Kang CK, Fukui T, Doi Y (1996) Production of a novel copolyester of 3-hydroxybutyric acid and medium-chain-length 3-hydroxyalkanoic acids by Pseudomonas sp. 61–3 from sugars. Appl Microbiol Biotechnol 45:363–370

    Article  CAS  Google Scholar 

  • Koller M (2019) Polyhydroxyalkanoate biosynthesis at the edge of water activity-Haloarchaea as biopolyester factories. Bioengineer 6:34

    Article  Google Scholar 

  • Koller M, Hesse P, Bona R, Kutschera C, Atlić A, Braunegg G (2007) Biosynthesis of high quality polyhydroxyalkanoate co- and terpolyesters for potential medical application by the archaeon Haloferax mediterranei. Macromol Symp 253:33–39

    Article  CAS  Google Scholar 

  • Koller M, Chiellini E, Braunegg G (2015) Study on the production and re-use of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) and extracellular polysaccharide by the archaeon Haloferax mediterranei strain DSM 1411. Chem Biochem Eng Q 29:87–98

    Article  CAS  Google Scholar 

  • Kusaka S, Iwata T, Doi Y (1998) Microbial synthesis and physical properties of ultra-high-molecular-weight poly[(R)-3-hydroxybutyrate]. J Macromol Sci Pure Appl Chem A 35:319–335

    Article  Google Scholar 

  • Lentzen G, Schwarz T (2006) Extremolytes: natural compounds from extremophiles for versatile applications. Appl Microbiol Biotechnol 72:623–634

    Article  CAS  PubMed  Google Scholar 

  • Liu C, Baffoe DK, Zhan Y, Zhang M, Li Y, Zhang G (2019) Halophile, an essential platform for bioproduction. J Microbiol Method 166:105704

    Article  CAS  Google Scholar 

  • Liu G, Cai S, Hou J, Zhao D, Han J, Zhou J, Xiang H (2016) Enoyl-CoA hydratase mediates polyhydroxyalkanoate mobilization in Haloferax mediterranei. Sci Rep 6:24015

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Liu G, Hou J, Cai S, Zhao D, Cai L, Han J, Zhou J, Xiang H (2015) A patatin-like protein associated with the polyhydroxyalkanoate (PHA) granules of Haloferax mediterranei acts as an efficient depolymerase in the degradation of native PHA. Appl Environ Microbiol 81:3029–3038

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lu QH, Han J, Zhou LG, Zhou J, Xiang H (2008) Genetic and biochemical characterization of the poly(3-hydroxybutyrate-co-3-hydroxyvalerate) synthase in Haloferax mediterranei. J Bacteriol 190:4173–4180

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • McChalicher CWJ, Srienc F (2007) Investigating the structure-property relationship of bacterial PHA block copolymers. J Biotechnol 132:296–302

    Article  CAS  PubMed  Google Scholar 

  • Melanie S, Winterburn JB, Devianto H (2018) Production of biopolymer polyhydroxyalkanoates (PHA) by extreme halophilic marine Archaea Haloferax mediterranei in medium with varying phosphorus concentration. J Eng Technol Sci 50:255–271

  • Meseguer I, Rodriguez-Valera F (1985) Production and purification of halocin H4. FEMS Microbiol Lett 28:177–182

    Article  CAS  Google Scholar 

  • Mitra R, Xu T, Xiang H, Han J (2020) Current developments on polyhydroxyalkanoates synthesis by using halophiles as a promising cell factory. Microb Cell Fact 19:86

    Article  PubMed  PubMed Central  Google Scholar 

  • Mokashe N, Chaudhari B, Patil U (2018) Operative utility of salt-stable proteases of halophilic and halotolerant bacteria in the biotechnology sector. Int J Biol Macromol 117:493–522

    Article  CAS  PubMed  Google Scholar 

  • Ng E-L, Lwanga EH, Eldridge SM, Johnston P, Hu H-W, Geissen V, Chen D (2018) An overview of microplastic and nanoplastic pollution in agroecosystems. Sci Total Environ 627:1377–1388

    Article  CAS  PubMed  Google Scholar 

  • Oren A, Hallsworth JE (2014) Microbial weeds in hypersaline habitats: the enigma of the weed-like Haloferax mediterranei. FEMS Microbiol Lett 359:134–142

    Article  CAS  PubMed  Google Scholar 

  • Pais J, Serafim LS, Freitas F, Reis MAM (2016) Conversion of cheese whey into poly(3-hydroxybutyrate- co-3-hydroxyvalerate) by Haloferax mediterranei. New Biotechnol 33:224–230

  • Parolis H, Parolis LA, Boán IF, Rodríguez-Valera F, Widmalm G, Manca MC, Jansson PE, Sutherland IW (1996) The structure of the exopolysaccharide produced by the halophilic archaeon Haloferax mediterranei strain R4 (ATCC 33500). Carbohydr Res 295:147–156

    Article  CAS  PubMed  Google Scholar 

  • Rodriguez-Valera F, Ruiz-Berraquero F, Ramos-Cormenzana A (1980) Isolation of extremely halophilic bacteria able to grow in defined inorganic media with single carbon sources. J Gen Microbiol 119:535–538

    Google Scholar 

  • Sasuga T, Hayakawa N, Yoshida K, Hagiwara M (1985) Degradation in tensile properties of aromatic polymers by electron beam irradiation. Polymer 26:1039–1045

    Article  CAS  Google Scholar 

  • Sim SJ, Snell KD, Hogan SA, Stubbe J, Rha CK, Sinskey AJ (1997) PHA synthase activity controls the molecular weight and polydispersity of polyhydroxybutyrate in vivo. Nat Biotechnol 15:63–67

    Article  CAS  PubMed  Google Scholar 

  • Spierling S, Knüpffer E, Behnsen H, Mudersbach M, Krieg H, Springer S, Albrecht S, Herrmann C, Endres H-J (2018) Bio-based plastics—a review of environmental, social and economic impact assessments. J Clean Prod 185:476–491

    Article  Google Scholar 

  • Sudesh K, Abe H, Doi Y (2000) Synthesis, structure, and properties of polyhydroxyalkanoates: biological polyesters. Prog Polym Sci 25:1503–1555

    Article  CAS  Google Scholar 

  • Tanadchangsaeng N, Kitagawa A, Yamamoto T, Abe H, Tsuge T (2009) Identification, biosynthesis, and characterization of polyhydroxyalkanoate copolymer consisting of 3-hydroxybutyrate and 3-hydroxy-4-methylvalerate. Biomacromolecules 10:2866–2874

    Article  CAS  Google Scholar 

  • Torregrosa-Crespo J, Martínez-Espinosa RM, Esclapez J, Bautista V, Pire C, Camacho M, Richardson DJ, Bonete MJ (2016) Anaerobic metabolism in Haloferax genus: denitrification as case of study. Adv Microb Physiol 68:41–85

    Article  CAS  PubMed  Google Scholar 

  • Tsuge T (2016) Fundamental factors determining the molecular weight of polyhydroxyalkanoate during biosynthesis. Polym J 48:1051–1057

    Article  CAS  Google Scholar 

  • Zuo Z-Q, Xue Q, Zhou J, Zhao D-H, Han J, Xiang H (2018) Engineering Haloferax mediterranei as an efficient platform for high level production of lycopene. Front Microbiol 9:2893

    Article  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

The authors thank Ms. Hiroko Shinozaki for technical assistance.

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SS conceived and designed research, conducted experiments, and drafted the manuscript. KI conducted experiments and drafted the manuscript. KU discussed the results and drafted the manuscript. KW, AS, TF, KO, and TM discussed the results. All authors read and approved the manuscript.

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Correspondence to Shun Sato.

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Sato, S., Ino, K., Ushimaru, K. et al. Evaluating haloarchaeal culture media for ultrahigh-molecular-weight polyhydroxyalkanoate biosynthesis by Haloferax mediterranei. Appl Microbiol Biotechnol 105, 6679–6689 (2021). https://doi.org/10.1007/s00253-021-11508-3

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